Role and Applications of Boron and Its Compounds in Biomedicine, Health and Agriculture

#### **Chapter 1**

## Physical, Biological, and Clinical Merits of High Energy Boron Ions for Radiation Therapy

*Anders Brahme*

#### **Abstract**

The lightest ions beyond protons, principally helium, lithium, and boron ions, make highly specific molecular Bragg peak radiation therapy of malignant tumors possible with minimal adverse normal tissue reactions. The Bragg peak ionization density is mainly elevated in a few mm wide spot at the end of these ions with substantially increased local apoptosis and senescence induction. Mainly placing Bragg peaks in the gross tumor volume with increased local therapeutic effect and only low ionization density and easily repairable damage in normal tissues. The possible geometrical accuracy of the dose delivery will be ≈1 mm with these ions. Interestingly, high-resolution molecular tumor imaging will then be possible, particularly with 8 Boron ions that are our lightest positron emitter allowing immediate accurate PET-CT imaging to delineate the target volume dose delivery. Compared to carbon ions the boron radiation damage to normal tissues in front of and behind the tumor is reduced at the same time as tumor apoptosis and senescence are increased. A mean tumor cure as high as 80% should be possible with Boron ion therapy using new clinical fractionation principles and even more when early tumor detection and malignancy estimation methods are brought into more regular clinical use.

**Keywords:** boron ion radiation therapy, 8−11boron ions, low dose apoptosis, low dose hypersensitivity, high dose apoptosis, radiation dose-response relationships, light ion radiation therapy, radiation therapy optimization

#### **1. Introduction**

The interaction of ionizing radiation with living tissues and tumors is one of the most complex biomedical problems, since it requires knowledge about atomic and nuclear physics and the generated secondary electrons, as well as the molecular biology dynamics of living tissues and cells and their complex damage repair systems [1]. Understanding radiation-induced cellular damage and repair is the key to optimal safety in the therapeutic and the diagnostic use of high-quality radiation beams. Biologically optimized intensity-modulated photons, electrons, and light ions represent the ultimate development of radiation therapy as shown in **Figure 1**. The absorbed dose and biological effect on normal tissues can be designed so it is as low as possible from a radiation physical point of view, at the same time, as the therapeutic

#### **Figure 1.**

*Illustration of the fantastic power available by using biologically optimized inverse radiation therapy planning [2]. If we know the approximate sensitivity of the tumor and the normal tissues (!), it is possible to derive the biologically optimal beam directions and their intensity modulation (?; [3]) and it is even possible to find the optimal combination of low and high ionization density radiations (cf.* **Figure 15***) and their incident energy spectra as well as the ideal time dose fractionation [1, 4, 5] using biological complication free cure (P+) optimization strategies (P++: P+ with concomitant injury minimization [3]). In addition, if we have information about the interaction of the radiation modality of interest with chemotherapeutic agents of preference, the combined treatment schedule can also be optimized in biological terms. During the last week of treatment, only a hand full of tumor clonogens remain, and should thus preferably be treated with more microscopically uniform electron or photon beams. This is optimal since both the particle beam and the tumor cells are quantized and may protect some tumor clonogens from lethal hits by inevitable cold spots between the ions during the last most curative therapeutic dose fraction [1–5]!.*

effect on radiation-resistant tumor cells is as high as possible from a radiation biological point of view [2]. With the lightest ions above protons: He, Li, B, and C the border region between the clinical gross tumor and target volume and surrounding healthy normal tissues can be set as narrow as physically possible. In addition, the optimal number of treatment fractions can be substantially reduced, and the curative gain factor on radiation resistant hypoxic tumor cells may generally be more than doubled compared to low ionization density photons, electrons, and protons. **Figure 1** shows how the optimal selection of Therapeutic beams can be arranged and the energy modulation shaped to maximize the cure probability of the patient with minimal risk for side effects in normal tissues [2]. Largely based on clinically established doseresponse parameters of normal tissues (*γ, D*50*, s*).

### **2. Radiation biology of radiation therapy**

#### **2.1 Handling of DNA damage by the TP53 gene**

The new interaction cross-section based Repairable-Homologous-Repairable damage formula for radiation-induced cellular inactivation, repair, misrepair, and apoptosis in TP53 intact and mutant cell lines can be used to optimize radiation therapy. The formulation requires renewed thinking about the biological

#### *Physical, Biological, and Clinical Merits of High Energy Boron Ions for Radiation Therapy DOI: http://dx.doi.org/10.5772/intechopen.111485*

optimization of radiation therapy. It suggests that most TP53 intact normal tissues are Low Dose Hyper Sensitive (LDHS, see left insert of **Figure 2**) and that the inherent microscopic heterogeneity of higher Linear Energy Transfer (*LET*) ion treatments the last week would benefit from low *LET* as shown in the lower right of **Figure 2** [1, 4, 5]. The ability of the new method to quantify apoptosis [1], has helped identify the early Low Dose Hypersensitivity (LDHS) and Low Dose Apoptosis (LDA) of most normal tissues and tumors with intact TP53 and ATM genes. This mechanism has probably been developed by nature's proses of survival advantage selection, to ensure minimal risk for severe mutations to the genome before the DNA repair system is fully functional after around a dose of 1Gy [1, 4–8]. As a compensating measure the apoptosis-inducing caspase 3 gene product (**Figure 2** right) remarkably "remembers" this low dose apoptotic cell loss and starts cellular repopulation to reestablish homeostasis in the tissues after being irradiated.

This useful mechanism in normal tissue is a well-known problem after suboptimal radiation therapy where it can cause accelerated repopulation of remaining tumor cells at the end of treatment [9]. A clear curative intent is probably the principal way to avoid this tumor-reactivating mechanism. These studies also identified that maximum apoptosis is induced by the lowest *LET* ions largely as they have the highest fluence of *δ*-electron apoptosis induced by primary ions per unit dose [1, 4]. With a too high *LET* the apoptosis and senescence will instead be high in the normal tissues in front of and behind the tumor, which may be undesirable from a complication-free

#### **Figure 2.**

*The complex responses of the TP53 gene to mild and severe genetic stress is largely determining the cellular response to radiation [1–13]. Mild stress phosphorylates the serine 15 and 20 sites on p53 by ATM and CHK2, resulting in cell cycle block and DNA repair. This results in LDHS in normal tissues but generally not in tumors often with a mutant TP53 gene as seen in the cell survival insert. Local high doses or high ionization densities resulting in DDSBs (Dual Double Strand Breaks cf.* **Figure 3***) [2, 14] are increasing the severity of the damage phosphorylating also the serine 46 site e.g., via DYRK2, p38K, or ATM, and a high dose apoptotic (HDA) response may get triggered. Lithium-Boron (cf* **Figure 12***) ions allow unique therapeutic use by inducing a massive apoptotic-senescent tumor cell response within the Bragg peak (σ*h *homologically repairable damage and σ*i *direct inactivation cross-sections cf.*  **Figure 4** *[1, 4]), but in front of and beyond the Bragg peak, the LET is low, and non-homological easily repairable damage is mainly induced (σ*n *cross-section see* **Figure 4** *insert [1: Figure 8, 4, 7, 8]).*

cure point of view, even if hypoxic tumors may marginally benefit from a high *LET* (cf [4] and **Figure 3**).

#### **2.2 Dual double strand breaks**

The severe Bragg peak damage to supercoiled DNA first wound two times around nucleosomes in the cell nucleus is shown in the lower right corner of **Figure 2** and the close-up in **Figure 3**. Most toxic are the ≈700 eV electrons that deposit a dose in the neighborhood of the track of up to 10<sup>6</sup> Gy as seen in the right of **Figure 2** [1, 4, 14, 15]. In front of the Bragg peak, the density of electrons is lower and even more so in the high energy entrance region requiring many more ions to deliver a dose of around 2Gy (cf **Figure 7**). This phenomenon explains why the medium and low *LET* ion beams are most efficient in inducing apoptosis (as further seen in **Figures 11** and **13**) and thereby eradicating hypoxic tumor cells. With a low *LET*, too few severe direct cell kill events are obtained and at high *LET* too few ions are available at a given dose even though they produce very severe damage. The most probable DNA fragment length at high doses is around 78 base pairs, corresponding to a single turn around the nucleosome (cf **Figure 3**). A δ-electron track end that may randomly hit the DNA at any point on the periphery of the nucleosome and then often produce a Dual DSB (DDSB), can easily produce such fragments. This will very often make DNA fragments of close to a single nucleosomal DNA turn in length as seen in **Figure 3**. This string of DNA may easily be lost as a micronuclei or gey inserted erroneously to make a severe mutation and possibly a non-functional protein. Fortunately, most simple DSBs are repaired

#### **Figure 3.**

*Molecular close-up of ion tracks (right) showing that most of the lethal cell damage of densely ionizing ions is induced by low energy δ-electrons in the 200 eV to 1 keV energy range generating severe difficult-to-repair DNA damage in the cell nucleus such as dual DSBs at the periphery of the nucleosome (left, cf. [14]). The left insert shows that the most common DNA segment length corresponds to a single turn of DNA around a nucleosome as expected from the DDSBs at the periphery of a nucleosome. Interestingly, the 78–80 base pair fragments are about twice as common as all other fragment sizes and they should be expected to be even more common with high LET beams having* ≈*3 times more secondary δ-electrons in the sub-keV energy range, with a very high probability of inducing lethal DDSBs.*

correctly (>99%) so a DDSB is really the most common multiply damaged site causing severe cell loss [1, 4, 14, 16].

#### **2.3 Cell survival**

In two recent DNA repair-based publications [1, 4], the accurate quantification of the cellular survival and damage to tumors and normal tissues are developed to significantly improve our ability to precisely describe the survival to low and high ionization density (*LET*) radiations and doses S = e<sup>−</sup>*aD* + *bD*e<sup>−</sup>*cD*, far beyond the possibilities of the conventional linear quadratic cell survival model (S≈ <sup>−</sup>α −β e *D D***<sup>2</sup>** ). Not only are the undamaged cells (e−aD) separated from the sublethal damaged cells (*bD*) but also the two major DNA damage repair pathways, namely homologous and non-homologous end joining DNA repair (HR, NHEJ) can each be identified

(*b*h*D*e<sup>−</sup>*c*h*<sup>D</sup>* + *b*n*D*e<sup>−</sup>*c*n*<sup>D</sup>* ≈*bD*e<sup>−</sup>*cD*) and so can their complex interactions cf.

#### **Figures 4** and **5** [1].

It is therefore given the name the repairable-homologous-repairable or RHRformulation as seen in the left insert in **Figures 2** and **4** (cf [1, 4] for further details). The fractionation window linked to LDHS normal tissues as seen in the left insert in **Figure 2** indicates that the low *LET* dose to organs at risk should be ≈2 Gy/Fr as this produces the least damage per unit dose, whereas the tumor dose should be substantially higher to ensure perfect tumor cure [5]. This calls for biologically optimized treatments using a few intensity-modulated beams [3] to avoid secondary cancer risk and get a true curative intent, avoiding caspase 3 induced accelerated tumor cell repopulation (see right part of the middle half of **Figure 2**, [9]). Light ions with the lowest possible *LET* in normal tissues and high *LET* only in the tumor indicate lithium to boron ions [5]. The high microscopic heterogeneity in the tumor will cause local microscopic cold spots. Therefore, the last week of curative ion therapy, with few remaining viable tumor clonogens randomly spread in the target volume, as indicated in **Figure 1**, should receive the last 10 GyEquivalent by low *LET* ensuring perfect microscopic tumor coverage and high cure and reduced risk for adverse reactions in normal tissues [5]. Interestingly, such an approach would also ensure a steeper rice of tumor cure and a higher complication-free cure as few remaining clonogens are fairly well oxygenated eliminating shallower tumor response by ion heterogeneity (see **Figure 17**). Avoiding ion microscopic heterogeneity in normal tissues increases complication-free cure both at the low dose normal tissue complication side and high dose tumor cure end of the treatment [5].

#### **2.4 Apoptosis induction**

The major forms of interaction between homologous and non-homologous DNA repair such as homologous repair of non-homologous misrepair are accounted for and the probability to induce programmed cell death (Apoptosis) and potentially even more so, permanent cell cycle arrest (Senescence) as well as other cell cycle losses [3, 4]. Interestingly, these processes are probably the optimal ways to inactivate a tumor with minimal inflammatory response and without massive immediate tumor decomposition. Interestingly, lithium ions and its neighbors helium, beryllium, and boron ions have an important and unique potential to induce apoptosis locally, mainly in a few mm-size volumes around their deep high ionization density Bragg peaks in the tumor as seen in **Figures 4, 5** and **7** (Be may be associated

#### **Figure 4.**

*The cell survival, the cell fractions that are totally un-hit by the beams, and the apoptotic and non-apoptotic death over the LET range 0.3-40-80-160 eV/nm from 60Co and boron ions. The cell survival shows a gradual increase in steepness with increasing LET whereas the Afr has its maximum at a dose causing around 13.5% cell survival as indicated by the arrows. For the two lowest LET's, the non-apoptotic cells, upper dashed curves, and the clonogenic survival are practically tangential at low doses indicating apoptosis is the preferred way of cell death before p53 is phosphorylated at its Serine 15 and 20 sites at* ≈*½ Gy (***Figure 2***). The shaded area is due to non-apoptotic cell death for 40 eV/nm boron ions showing the domination of apoptotic cell death at the low LET's as the shading is lost at low doses. The survival data are also used in* **Figure 14** *on a linear scale to derive the secondary cancer induction probability. The associated LET variation of the non-homological and homological interaction cross-sections n and h for DNA repair after 10B irradiation but also for 12C ions, as shown by the insert [1, 4]. A comparison with the variation of the fast b*n *and slow b*h *repair of repair some foci after 14N irradiation (dashed lines, right scale, data derived from [4, 17]). The homological cross-section h increases very fast with the LET for 10B ions due to rapidly narrowing δ-electron cores and so is the associated reduction of the n cross-sections.*

#### **Figure 5.**

*The LET variation of the apoptotic fraction contributions of the eight key misrepair processes A-N indicating a relative apoptotic effectiveness (RAE) of about 3.4 for low LET boron ions around 40 eV/nm, whereas the peak relative biologic effectiveness (RBE) is about 3.5 but closer to an LET of about 140 eV/nm (Modified from [1]).*

*Physical, Biological, and Clinical Merits of High Energy Boron Ions for Radiation Therapy DOI: http://dx.doi.org/10.5772/intechopen.111485*

with additional BeO forming toxicity in the tumor but also in normal tissues cf. **Figures 11** and **13**). Everywhere else they mainly induce low-ionization density and LET-type effective DNA repair processes as seen in the right part of **Figure 2**. The surprisingly popular proton therapy has an almost negligible such augmented local high biological tumor effect, helium has some, lithium, beryllium, and boron have more and more, and finally the already rather promising carbon ions have a little too much, both in the entrance region with mainly normal tissues and the fragmentation tail beyond the tumor, and not least up to as far as 5 cm in front of the Bragg peak (cf **Figure 12**). Boron ions are therefore most likely the optimal ion for medium to large tumors whereas for small to medium size tumors and not least pediatric tumors a combination with lithium ions may be the most optimal for clinical use (cf **Figures 2** and **15**)! Interestingly, the lightest existing positron emitter Boron 8, makes it possible to immediately visualize the delivered dose to the patient by Positron Emission Tomography (PET) and verify that optimal dose delivery is achieved in the tumor region.

#### **2.5 Relative biological effectiveness**

With increasing atomic number, the penumbra gets narrower and the longitudinal range straggling is also lower, so more of the energy is deposited in the tumor by light ions of increasing atomic weight and Relative Biological Effectiveness (*RBE*) as seen in **Figure 6**. The high energy deposition density at the end of the ion range (the Bragg peak) is caused by a velocity resonance increasing the energy transfer to the tumor cells when the speed of the ion is close to the speeds of the orbital electrons of the tumor tissue and there is an increased probability for high energy transferred from the multiply charged ions to the electrons as they travel longer distances together toward the end of the particle range. The resulting peak is seen in **Figures 2, 6–8, 11** and **12**. When the atomic weight gets too high, the amount of particle fragments increases, so the dose beyond the Bragg peak gets high too, as seen in **Figures 12** and **13**. Ions heavier than carbon should therefore be used very carefully with sensitive normal tissues in front of and beyond the tumor.

#### **3. Physical, biological, and clinical properties of light ion beams**

Compared to low *LET* photon, electron, and proton beams, the clinical properties of light ion beams for radiation therapy are much more versatile and complex as discussed in more detail below. A large part of the detailed specific information is presented in graphical form in the Figures and their captions for simplicity and clarity.

#### **3.1 Ion pencil beams**

The physical and biological properties of narrow beams of the six lightest ions when penetrating water to a depth of 30 cm by their central axis energy deposition density profiles already shown in the lower row of **Figure 1** in 2D are described in 3D. The influence of multiple Columb scattering and longitudinal range straggling on the Bragg peak of the pencil beams is clearly shown in **Figure 7** (see also **Figures 8–10** below). With protons, the dose to normal tissues in front of the tumor is twice the

#### **Figure 6.**

*Comparison of the RBE and LET ranges available with protons, lithium, boron, carbon, and neon ions is shown. It is seen that the range from lithium to carbon ions is most interesting, especially for hypoxic tumors. In general, LETs beyond the RBE peak should be avoided to minimize normal tissue damage in the entrance and plateau region (C-Ne, cf.* **Figure 2***). As the cross-section saturates (cf insert in* **Figure 3***), the relative biological effectiveness RBE reaches a maximum since the cross-section cannot increase with the LET anymore, and at higher LETs the biological effectiveness decreases because of an increased probability of radical—radical recombination as secondary electrons are generated more and more closely together. Furthermore, the "overkill" effect implies that multiple kill events are equal to a single kill (you can only die once). The dashed and solid curves [18] describe the average response of the multiple experimental data sets very well [19].*

tumor dose due to significant multiple scatter whereas it is only a small fraction of the tumor dose for the light ions of lithium, boron, and carbon. This makes the local increase in dose to a small region of normal tissues in front of the tumor about five times larger than the increase in tumor dose that may be needed for tumor cure. This demonstrating that for each dose addition to a small part of a tumor volume will require that about five times more doses have to be given to normal tissues in front of the tumor with protons as compared to other light ions from lithium to boron. This is a severe dose delivery disadvantage for small radiation-resistant tumors and extra beam portals may be needed to avoid normal tissue damage with protons. It may be less of a dosimetric problem for large tumors where the broad beam Bragg peak dose level is better established. However, large tumors often have extensive hypoxic regions so protons are not generally the radiation modality of choice, and lithium to boron and even carbon ions are more often indicated for larger hypoxic tumors. The shapes of the so-called narrow pencil beams in **Figure 7**, therefore, are a kind of figure of merit when using inverse biologically optimized treatment planning trying to maximize the patient fraction that is cured without severe damage to normal tissues [3]. A high distal the so-called Bragg peak is therefore important and so is a low dose in the entrance and exit regions to minimize normal tissue damage in front of and not least behind the tumor by nuclear fragmentation as seen in even more detail for Boron and Carbon in **Figure 12**.

*Physical, Biological, and Clinical Merits of High Energy Boron Ions for Radiation Therapy DOI: http://dx.doi.org/10.5772/intechopen.111485*

#### **Figure 7.**

*3D Illustration of the clinical value of different 5 mm 1/e width light ion pencil beams for biologically optimized therapy planning. For carbon ions and heavier, the increasing LET in the plateau-type entrance region has to be considered more carefully when maximizing the probability of reaching a complication-free cure. The color scale illustrates the ionization density (LET) and consequently the increased biological effect in the tumor, which comes as a very important biological advantage on top of the physical dose distributional advantage shown in the figure.*

#### **Figure 8.**

*As the cross-section increases with nuclear size light ion attenuation will be less in the normal tissues of the patient, to more effectively reach a deep sited tumor! Hydrogen would thus be optimal from this point of view but the biological effect is minimal as seen in* **Figures 2** *and* **4***, making He, Li, and B more interesting!.*

#### **3.2 Exponential ion attenuation**

Like Photons, Light ions are attenuated exponentially but by nuclear reactions rather than by photoelectric, Compton, pair, and photonuclear reactions. Protons are only weekly attenuated, less than 50 MV photons, whereas carbon ions are similarly attenuated as 16 MV photons and neon more like 4MV and almost 60Co. Part of the clinical problem with Neon ions is very clear as they are severely attenuated at large depths. **Figure 6** shows the other part of their problem as their entrance regions have a very high RBE and they reach far into the overkill region at depth as also seen in **Figure 11**. The merits of the light ions from lithium to boron are clearly shown in the figure all with an attenuation less than that of a 45 MV Betatron beam. Furthermore, to reach 25 cm of tumor depth a 400 MeV/u carbon ion beam may be needed whereas 300 MeV/u boron may suffice, thus requiring a smaller and less costly cyclotron for beam production as seen in the figure.

#### **3.3 Particle multiple columb scattering**

The increase in the penumbra (distance between 80 and 20% isodose lines) as a function of depth in water is shown in **Figures 9** and **10**. The Multiple Columb Scattering in the patient will increase the penumbra substantially with depth since the scattering power increase with decreasing energy. On top of these, multiple scatter contributions (*r*1/*e* = σ*r*) the part from the initial effective source size (σ0) of the intrinsic accelerator beam should be added in quadrature as they stem from statistically independent processes so the total standard deviation is given

#### **Figure 9.**

*With electrons and protons, the penumbra width is better than for photons at shallow depths (<5 cm), whereas light ions from helium and beyond are needed to get significant improvements compared to photons at large tumor depths. The insert shows the clinical advantage of a sharp penumbra in the neighborhood of organs at risk. The brainstem in this case is almost totally avoided with carbon or boron ions but not with protons (courtesy Jürgen Debus, Heidelberg).*

*Physical, Biological, and Clinical Merits of High Energy Boron Ions for Radiation Therapy DOI: http://dx.doi.org/10.5772/intechopen.111485*

#### **Figure 10.**

*The variation of the lateral penumbra and longitudinal Bragg peak width and range straggling for light ions of increasing nucleon number. A sharp reduction to 50% of the wide half-value width of protons is seen for helium and about one-third for lithium and about a quarter for boron and beyond.*

by σtot<sup>2</sup> = σ<sup>r</sup> 2 + σ<sup>0</sup> 2 . This initial part is included in **Figure 7**, where σ0 = 2,5 mm (1/*e* width = 5 mm). The clear improvement in tumor coverage and normal tissue avoidance is seen in **Figures 9** and **10**, where also the reduction in penumbra width is seen beyond helium ions. In **Figure 10** the initial very steep reduction in the lateral penumbra and the longitudinal range straggling when going from protons to helium and more gradual going from helium to lithium, boron and carbon is clearly demonstrated. Since the penumbra of helium ions (α-particles) is already half of that for protons, helium ions are really the particle of choice in the low *LET* region as they in addition have very clear biological advantages not least in hypoxic tumors (cf **Figures 2, 6, 7, 9** and **11**).

#### **3.4 Lateral penumbra width and longitudinal range straggling**

The lateral multiple scattering and the penumbra decrease in almost the same fashion as the longitudinal straggling measured at the 60% width of the high *LET* region of the Bragg peak and they are closely proportional to the inverse square root of the nucleon number as seen **Figure 10**. The full 60% Bragg peak width is approximately the half width of these former quantities as seen in the right insert [20]. One may ask why there is such a large difference in biological effect between protons and other light ions (see **Figure 7**) even though their normalized broad beam dose distributions are fairly similar (cf **Figures 8** and **11**). This is mainly due to the same phenomenon that reduces their penumbra width and longitudinal range straggling with increasing nucleon number as seen in the middle straggling insert in **Figure 10**, reducing the single track LET from about 70 eV/nm over some 10 *μ*m to a mean value of about 5 eV/nm in the few mm of range straggling at the Bragg peaks in clinical beams. This also shows up in the small *RBE* variation of protons in **Figure 11**.

#### **Figure 11.**

*The Variation of the biological effect over the SOBP will make the dose at the distal target volume low and LET high making the risk for microscopic cold spots high and increasing the risk for a recurrent tumor [5]. With high energy electrons and photons two perpendicular beams make a better high-dose dose distribution than a proton SOBP even if the low dose is lower, making Li, B, and C ions most interesting from a therapeutic point of view. With B and C ions, the method with two different intensity-modulated beams will eliminate the single beam SOBP problem as shown in* **Figure 15***.*

#### **3.5 The** *RBE* **variation in the beam**

The traditional way to make a uniform dose in an extended tumor volume is to use the so-called Spread Out Bragg Peak (SOBP) method first developed for protons at Berkeley and Uppsala where the energy and thus the range was modulated to get a uniform dose in the tumor volume [21]. This works well for protons with almost negligible biological effect variation with depth as seen in **Figure 11**. However, with Boron and Carbon ions there will be a substantial RBE variation with depth so the distal dose will be about half of that at the anterior part of the tumor generating an approximately uniform mean cell kill but a about twofold variation in biological effect as seen in **Figure 11**. This is far from desirable for a uniform tumor or even a heterogeneous tumor as seen from the large RBE variation measured for carbon ions in **Figure 11**. Combining lithium and boron ions in suitable ratios is better (**Figure 15**).

#### **3.6 LET variation in beams**

To clearly show the physical and biological differences between boron and carbon ion beams their partial ionization density contributions along with their absorbed dose distributions are shown in **Figure 12**. Even if their physical dose distributions are quite similar as already seen in **Figure 7**, their local ionization densities and LET's are rather different with the whole entrance region of carbon being of medium *LET*. With boron, this region is mainly low *LET* and the high *LET* region only extends ≈2 cm in front of the Bragg peak whereas it is about 5 cm for carbon ions. Furthermore, there is negligible elevated *LET* behind the Bragg peak of boron, so normal tissues both in front of and behind the tumor are much less damaged by boron therapy which is an

*Physical, Biological, and Clinical Merits of High Energy Boron Ions for Radiation Therapy DOI: http://dx.doi.org/10.5772/intechopen.111485*

#### **Figure 12.**

*Carbon and boron ion beam depth distribution of the energy deposition density. It is clearly seen that the adverse Biological effect in the entrance and fragmentation tail regions are significantly reduced by boron ions. With sensitive organs at risk in front of and behind the tumor volume, the high LET reduction will also be a further important advantage of boron ions as they will generate more apoptosis in the tumor than carbon ions as seen in*  **Figure 16***.*

#### **Figure 13.**

*Ion absorbed dose per neutron energy fluence. The increase of the RBE, the Bragg peak and fragmentation tail doses, DBp and DF per unit neutron energy fluence, ΨN, and ion fluence, ΦI, at tumor lethal doses of light ions as a function of the mean Bragg peak LET values are shown. All the values are given relative to those for protons and the curves clearly show how much higher the fluence of protons is compared to other light ions up to neon at a given biological effect. Furthermore, the increase in therapeutically effective dose in the Bragg peak per generated unit of neutron energy fluence, the ratios of tail fragment dose compared to the Bragg peak dose, the Bragg peak absorbed dose per neutron energy fluence, the increasing variance of the particle fluence as caused by the reduced fluence of ions at high LET s and RBEs, and the mean ion Bragg peak RBE are all shown in order from top to bottom.*

important clinical advantage! This makes boron ions most suitable for mixed beam therapy as is more clearly shown in **Figure 15**.

#### **3.7 Neutron production**

Like high-energy photon beams light ion beams are always associated with a significant neutron production as seen in **Figure 13**. Due to nuclear reactions, the cross-section and number of neutrons produced increases with the increasing atomic number of the projectile as the number of fragmentation reactions and consequently, the number of neutrons increases with nuclear size. However, the number of ions needed to eradicate a hypoxic tumor simultaneously decreases rapidly since the *LET* and *RBE* increase with atomic charge and mass as seen in **Figures 6** and **13**. In fact, **Figure 13** shows that the fluence reduction increases both the dose and the equivalent dose per unit neutron energy fluence generated and thus steadily decreases the neutron production with increasing atomic number of the ion at tumor lethal doses. Interestingly, at the same degree of cell kill, protons produce the highest neutron energy fluence partly because their mass is close to that of the neutron. From beryllium and above, the fragmentation is rather high so helium and lithium are probably the most ideal ion species for the treatment of pediatric malignancies. From the point of view of neutron and light fragment production, the ideal ion is generally somewhere between lithium and boron as seen in **Figure 13** but avoiding beryllium's high fragmentation and potential toxicity. Interestingly, even if the cross-section per ion to produce neutrons goes up slowly with atomic number, about 5, 15, and 40 times higher fluencies of protons are needed compared to helium, lithium, and carbon ions, making the total neutron production the highest by protons at equally curative doses of light ion therapy. Both the mean absorbed

#### **Figure 14.**

*The probability for secondary cancer induction as a function of the dose delivered to normal tissues. At low doses the risk of inducing a mutation is small, at high doses the probability to generate it is higher but so is the probability to eliminate it by the treatment. The risk is high in normal tissues between 0.5 and 6 Gy so this volume in the patient should be minimal! The LDA and LDHS of this TP53 intact cell line are clear! Interestingly, the risk is the smallest for the lowest LET boron ions due to HDA! CDN1: One-dimensional closest distance norm (not least square!). Based on data from [1, 5].*

*Physical, Biological, and Clinical Merits of High Energy Boron Ions for Radiation Therapy DOI: http://dx.doi.org/10.5772/intechopen.111485*

#### **Figure 15.**

*A quasi-uniform absorbed dose and cell kill distribution is generated between about 21 and 26 cm of depth by combining lithium and boron or carbon ions in suitable ratios to make the cell kill and survival quasi-uniform. The small local fluctuations in absorbed dose are due to a somewhat too large longitudinal range modulation (~3 mm) used to clearly illustrate the applied mechanism combining lithium and boron or carbon ion Bragg peaks at each depth interval. The different panels show the total absorbed dose and the boron or carbon doses and the lithium dose in the upper row, whereas the cell survival and mean LET distribution is shown below. Interestingly, by combining lithium and boron or carbon, a uniform biological effect, survival, and absorbed dose can be obtained both for uniform tumors and optimal biological effect modulation for heterogeneous tumors [23].*

dose (*D*Bp) and dose equivalent (*D*Eq,Bp) at the Bragg peak per unit energy fluence of neutrons generated (*Ψ*N) increase rapidly with the atomic number as seen in **Figure 13**, indicating that the absolute neutron production reduces steadily with atomic number. The dose of fragments is unfortunately also increasing. Helium and lithium ions are therefore indicated to be most optimal for pediatric tumors where the neutron fluence, dose and biological effect on normal tissues surrounding the tumor should really be minimized to reduce as far as possible the neutron-generated risk for secondary cancers.

#### **3.8 Secondary cancer induction**

To further illustrate the power of quantifying apoptosis, **Figure 14** estimates the probability of inducing secondary cancer based on experimental cell survival and apoptosis data [1: Figures 7+9]. It is unlikely that the Apoptotic fraction will contribute to secondary cancer induction (except possibly in TP53 mutant cell lines) so it is useful that this fraction can be estimated by the new RHR formula and be removed from other forms of misrepair to more accurately describe the cells that are potentially capable to generating secondary cancers. This cell fraction, as seen in **Figure 4**, has its peak in the 1-3 Gy range so in radiation therapy optimization, it is desirable to minimize this volume as much as possible in normal tissues. **Figure 14** show the maximal risk is smallest for low LET ions (blue shaded), the real risk may be in the order of 5% of the values in the Figure. This effect is a contraindication for very many beam portals in intensity modulated radiation therapy e.g., using "rapidarc", "volumetric

arc", and "tomotherapy" like methods on non-seniors, that may have time to develop secondary cancer after some 20 years cf [22].

### **4. Treatment optimization**

### **4.1 Generation of uniform absorbed dose and cell kill**

It is better from a microdosimetric point of view to generate a rather uniform microscopic energy deposition density on the cellular scale in the tumor to avoid dosimetric cold spots in tumor clonogens [5] that could repopulate the tumor and start with a slightly lower *LET* at the anterior tumor edge as shown in **Figure 15**. This could in principle be done in two different ways, either starting with a suitable Bragg peak at the distal tumor edge and gradually increasing the atomic weight and Bragg peak *LET* of the ion used or by just mixing two different ions species so that the mean *LET* and dose stays approximately constant as shown in **Figure 15**. Interestingly, this latter approach requires that most of the Bragg peak dose in the anterior part of the tumor is of very high *LET*, such as boron or carbon ions, whereas the distal part mainly requires a lower *LET* such as helium or lithium ions. In fact, mixing lithium and boron or carbon ions may be the optimal way to achieve close to ideal microscopic energy deposition density distribution for medium-to-large size tumors whereas several small oligo-metastasis are best treated by lithium ions alone. The survival was calculated for simultaneous irradiation whereas the mean *LET* is based both on lithium and boron or carbon ion components, but the *LET* variation was low, and

#### **Figure 16.**

*The LET variation of key biological parameters that influence the clinical value of radiation beams showing that the optimal window of opportunity in radiation therapy optimization is located between about 15 and 55 eV/nm or He-B ions. The underlying data are collected from Berkeley, NIRS, and Karolinska.*

*Physical, Biological, and Clinical Merits of High Energy Boron Ions for Radiation Therapy DOI: http://dx.doi.org/10.5772/intechopen.111485*

it peaks just downstream from the distal part of the tumor where the dose is a little low. The figure also illustrates the principle of Quality Modulated Radiation Therapy (QMRT) where the absorbed dose and biological effect and radiation quality can be optimally modulated instead of just the Intensity (IMRT) e.g., over a severely hypoxic tumor [23, 24].

#### **4.2 Selection of optimal particle and LET**

Based on the cell survival curves in **Figure 4** and apoptosis curves in **Figure 5**, one may ask which ion and LET should be used to maximize the complication-free cure of the radiation treatment. The very flexible method of mixing two ions with largely different ionization densities as shown in **Figure 15**, such as He or Li and B or C, rather than using Spread-Out Bragg Peaks [21], which work well only for protons as seen in **Figure 11**. The SOBP method generates strong variations in ionization density and absorbed dose. The variation is a factor of approximately two over the whole target volume for carbon ions, with a too low LET at the anterior end and too low dose and too high LET at the distal part of the target volume as seen in **Figure 11**. This significantly increases the risk of microscopic cold spots at the distal part of the target volume [5]. The use of mixed-modality treatments can make the absorbed dose and biological effect almost constant (cf **Figure 15**), or even locally elevated, e.g., in regions of strong hypoxia [2, 23, 24]. The beam quality question will therefore be discussed in terms of the optimal particle species and mean LET, as shown in **Figure 16** for the LET range from 0.2 to 180 eV/nm covering X-rays-Ne ions as recently discussed [4]. "In this region the RBE increases steadily from 1 to approximately 4.5, the oxygen enhancement ratio (OER) decreases from approximately 3 to almost 1, the normal tissue repair potential *b*/2*c* decreases from about 1 to ¼, the normalized clinical dose response gradient, *γ*C, decreases from approximately 6 to 2 (cf **Figure 17**), the microscopic standard deviation in dose *σ*μ increases from 1% to approximately 20%, the 50% tumor control dose, *D*50 decreases from almost 100 to 20 Gy for radiation-resistant tumors, the apoptotic fraction, *A*fr varies from almost 60 down to ≈3% and the oxygen gain factor (OGF) increases from 0 to 2.5. Importantly, above approximately 55 eV/nm, many of these factors become less advantageous for clinical use: the loss of sublethal DNA repair in normal tissues (see **Figure 4** with insert), the saturation of the OER and the OGF, and the reduced senescence and apoptosis in the tumor (**Figure 16**)". The increase in microscopic standard deviation (*σ*μ) will decrease the clinical *γ*C value and microscopic cold spots may appear as the standard deviation in dose delivery becomes more significant, as the therapeutic dose reaches ≈35 Gy and lower [5]. Consequentially, the senescence and apoptosis in the tumor decrease while it increases too much in normal tissues! Therefore, boron ions are more optimal than carbon ions (**Figure 12**), at least for medium-sized tumors, and lithium ions are the optimal particle for pediatric tumors, and their combination is ideal both for dose and radiation therapy biological effect optimization (**Figures 12** and **15**). It is very interesting to see that the apoptotic and senescent cell fractions have their maxima in the tumor at a much lower LET than the RBE, and it reaches approximately 60% at approximately 30 eV/ nm for boron ions (dotted curve in **Figure 14**), whereas the RBE has its maximum at around 140 eV/nm. This is mainly due to the larger number of ions needed per unit dose compared to carbon ions (cf. **Figure 13**). Carbon ions reach only approximately 40% at approximately 80 eV/nm and they may produce unnecessarily high senescence, apoptosis, and LET in normal tissues [4] as seen in **Figure 6**. Optimal

#### **Figure 17.**

*Microdosimetry of tumor control. Description of the reduction in the normalized steepness, γC, of the shape of the tumor control curve for uniform cell line for the different microscopic standard deviation of radiation modalities as a function of the absorbed dose (upper scale) and approximately normalized to the 50% tumor control dose (*≈*Dose Eq, lower scale, dashed lines) to more clearly see the effect on the γC value as the microdosimetric relative standard deviation increases with the LET. Not only do the hot spots often in the form of dual double strand brakes (DDSB, cf.* **Figures 2** *and* **3***) and cold regions get more extreme with increasing LET, but also the RBE is increasing, reducing the total dose about threefold with carbon, neutron, and neon, increasing the relative standard deviation and reducing the γC value more than one would like.*

cancer cell inactivation requires a treatment modality that preferentially induces senescence probably the most cost-efficient treatment to stop further cell cycling and block tumor growth. Actually, this is probably the mildest but still efficient endpoint to cure cancer [5, 10, 11, 25, 26], as it can make the clonogenic tumor cells lose their uncontrolled cell cycling ability as seen in **Figure 18**. The induction of autophagy (self-digestion) and apoptosis (programmed cell death, see **Figure 2**) may follow more severe DNA damage to minimize the cancer induction with more complex DNA damage and increased risk for severe mutations, e.g., with turnedon oncogenes or lost suppressor genes. Apoptosis and, even more so, senescence is probably the most optimal way to inactivate a tumor with minimal inflammatory response and without massive immediate apoptotic tumor decomposition [5, 25, 26]. Lithium ions, and its neighbors helium and boron, have an important and unique potential to induce apoptosis locally, mainly in a few mm-size volumes around their Bragg peaks in the tumor, and everywhere else, induce low-*LET* DNA repair facilitating molecular radiation therapy (see **Figure 2**). In repair terms, this means that NHEJ will dominate in the entrance and fragmentation tail of the ion, whereas HR will mainly try to handle the partly irreparable damage at the Bragg peak, which therefore should solely be placed into the tumor volume [1]. Thus, lithium ions are probably the optimal ion, at least for smaller tumors (see **Figures 2** and **11**, [10]) and pediatric patients. The major advantage of lithium ions is the low-ionization density in all normal tissues, largely inducing fast DNA repair while often inducing apoptosis and senescence only in the tumor volume. For the same reasons, medium size

*Physical, Biological, and Clinical Merits of High Energy Boron Ions for Radiation Therapy DOI: http://dx.doi.org/10.5772/intechopen.111485*

#### **Figure 18.**

*Illustration of the dose delivery for a large pelvic chordoma as shown in the upper and lower left treatment plans. The lower right plain MR images show the gradual disappearance of the chordoma 6 years after the treatment, probably due to a totally senescent response without traces of tumor growth. This treatment was made with carbon ions but could most likely be done with an advantage with boron ions as seen by the B and C ion LET data and apoptosis in* **Figures 7–17***! (Courtesy: Hirohiko Tsujii, NIRS, Chiba, Japan).*

tumors are probably best treated using boron ions, but this may sometimes require an extra beam portal compared to carbon ions, especially for larger hypoxic tumors. Interestingly, for radiation resistant commonly TP53 mutant tumors (cf **Figure 2** lower left insert) the p53 reactivating compound APR-246 can increase the apoptoticsenescent response and increase the effect of Reactive Oxygen Species (1: Figures 6+17, [5, 25, 27]) and will work well adjuvant also with Boron-ions. Furthermore, <sup>8</sup> B is our lightest and best direct PET emitter for concomitant online dose delivery imaging, although perhaps a little complex to make [28], and with a very short half-life of ≈0.8 sec so the PET camera really needs to be in the treatment room and used between ion accelerator pulses while treating the patient with the advantage of a rapid malfunction response time (10, 29, Figure 8.37, 8.38f).

#### **4.3 Influence microdosimetric heterogeneity on tumor control**

The shape of the Tumor Control curve is shown in **Figure 16** for a uniform cell line with different radiation modalities as a function of the absorbed dose (upper scale) and approximately normalized to the 50% tumor control dose (≈Dose Equivalent, lower scale) to more clearly see the effect of increasing microscopic heterogeneity as measured by the microdosimetric relative standard deviation (*σ*μ) with increasing ion mass, *LET* and *RBE.* Not only do the hot spots often in the form of Dual Double Strand Brakes (DDSB's, cf. **Figure 18**, [4: **Figure 2**]) and cold regions get more extreme with increasing *LET*, but also the *RBE* is increasing so the reduction of the total dose is about threefold with carbon, neutron, and neon increasing the relative standard deviation and reducing the clinical dose-response slope *γ*C severely. A steeper tumor control curve generally increases the therapeutic window of radiation therapy since the absorbed dose distribution and the associated therapeutic effect over the therapeutic window can be modulated with greater efficiency with a

steeper tumor cure and normal tissue damage curves [5]. Also with boron ions the last ≈10 GyE should therefore be delivered by low LET beams [5].

#### **4.4 Apoptosis and senescence**

Since apoptosis is nature's way to eliminate unwanted cells during the development of practically all organs and is therefore not generally associated with any inflammatory responses accompanying the more common necrotic type of cell kill. In addition, permanent cell cycle arrest or senescence and apoptosis are increasingly induced by light ions. Apoptosis through caspase 3 may cause accelerated tumor cell repopulation after a non-curative treatment [9]. Senescence may therefore be the most desirable endpoint of cancer therapy as the tumor cells lose their reproductive ability and are then slowly disappearing depending on the remaining cellular lifetime or half-life ≈2.5 years. As seen in **Figure 18** the tumor was reduced to about half the diameter or 10–15% of the initial volume 6 years after the treatment. Due to the fact that more ions per unit dose and cell kill are needed at medium to low ionization density, the more effective apoptotic and senescent response is obtained at an ionization density of around 20 eV/nm to 40 eV/nm as shown theoretically and experimentally in **Figures 15** and **16** [29]. Interestingly, helium, lithium and boron combine a high local apoptotic and senescent tumor cell inactivation only a few mm around their Bragg peaks and can thus be regarded as the ultimate stereotactic and conformal radiation modality (cf **Figures 1, 7, 12** [29–33]).

#### **5. Conclusions**

With boron ions the clinical experience from NIRS (National Institute for Radiological Sciences, Chiba) and other centers in Japan and Germany with carbon is an invaluable asset. A very promising development seen at NIRS is the very effective sterilization of large tumor masses (10–15 cm) such as pelvic chordomas as shown in **Figure 18** [29, 31–35]. These tumors show a gradual disappearance down to 10% and below the initial volume about 5 years after the treatment, probably due to a persistent senescent response likely to be even more effective with boron ions [35], and without evidence of any further tumor growth. The clinical value of light ion beams is discussed in further detail in a number of recent references [2, 5, 10, 19, 20, 29–34, 36]. When the above methods are brought into clinical use a mean tumor cure as high as 80% should be possible, and even more if the new therapeutic dose delivery principles [5], advanced dose fractionation [4: Figure 21, 5], and early tumor detection [10] and malignancy determination methods [37] come into more regular use.

*Physical, Biological, and Clinical Merits of High Energy Boron Ions for Radiation Therapy DOI: http://dx.doi.org/10.5772/intechopen.111485*

### **Author details**

Anders Brahme Department of Oncology-Pathology, Karolinska Institutet, Stockholm, Sweden

\*Address all correspondence to: andersbrah@gmail.com

© 2023 The Author(s). Licensee IntechOpen. This chapter is distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/3.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.

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#### **Chapter 2**

## Boron Removal by Donnan Dialysis According Doehlert Experimental Design

*Ikhlass Marzouk Trifi, Lasâad Dammak, Lassaad Baklouti and Béchir Hamrouni*

#### **Abstract**

Donnan dialysis is one of the membrane processes. It is based on the crossexchange of ions having the same electric charge through an ion-exchange membrane. The removal of boron by Donnan dialysis was studied in this work. First, a preliminary study was conducted to determine the experimental field of operating parameters using two membranes (AFN and ACS). Then, a full factorial design was applied to investigate the influence of the operating parameters and their interactions on the boron removal. Response surface methodology using Doehlert design was adopted to predict the optimal conditions. This approach via experimental designs is more efficient than the conventional optimization approach (the "one-at-a-time" method) which is time-consuming and requires a large number of experiments.

**Keywords:** boron removal, Donnan dialysis, response surface methodology, experimental design, anionic exchange membranes

#### **1. Introduction**

Sources of boron in the environment are mainly natural or even anthropogenic. It can be found mainly in the form of boric acid or borate salts. The presence of boron compounds in water increases in a continuous and parallel way to industrial development. In fact, boron is used in a wide range of industrial applications such as glass and ceramic industry to produce borosilicate glass, insulation, fiberglass, and flameretardant fiberglass [1]. Due to its high concentration of boron, natural water is often unsuitable for human consumption or agricultural use. Therefore, the harmful effects of boron on living organisms also increase, especially on plants, since boron manifests an important micronutrient-toxic boron duality [1]. Moreover, boron is a unique micronutrient in which overdose and underdose of boron supply cause toxicity and deficiency symptoms in plants, respectively. The level of boron in irrigation water exceeding 1 mg/L can affect the yield of sensitive crops (e.g., avocado and citrus fruits) [1]. Irrigation water with a very low boron content is required for certain metabolic activities, but when boron concentrations are increased to 4 mg/L [1], plants become poisoned, manifesting as yellow spots on leaves and fruits, and their

decomposition speeds up and they die [2]. According to the World Health Organization (WHO), drinking water should have a boron content of less than 0.3 mg/L because humans may also be poisoned by excessive boron levels [3, 4]. Consequently, several studies have focused on boron removal such as adsorption [5, 6], electrocoagulation [7, 8], or membrane processes namely electrodialysis [9, 10], reverse osmosis [11, 12], nanofiltration [13, 14], microfiltration [15], ion-exchange [15, 16], membrane distillation [17], and Donnan dialysis which was the subject of this study [18, 19]. This membrane process referring to FG Donnan [20] uses ion-exchange membranes allowing cross-exchange of ions to separate or concentrate ionic species. Modern Donnan dialysis uses ion-exchange membranes to separate the two solutions that are involved in the transport process. With no external electric potential difference applied across the membrane, Donnan dialysis uses an ion-exchange membrane. Donnan dialysis uses the counterdiffusion of two or more ions through an ion-exchange membrane to achieve separation. Donnan dialysis is one of the easiest and most inexpensive membrane techniques. The chemical potential gradient between the constituents of two solutions separated by a membrane is what propels the process. When Donnan equilibrium is reached, the process of Donnan dialysis is complete. It involves the stoichiometric exchange of counterions, or ions with the same charge, over an ion-exchange membrane [21].

There is frequently no theoretical model or one that is quite sophisticated that links certain controllable variables (factors) to a response. In this situation, empirical data should be used to determine the link between the causes and the response. Box and Wilson first presented the Response Surface Methodology (RSM), a group of mathematical and statistical tools whose goal is to evaluate situations like the one given using an empirical model. The RSM was used to explore the removal of boron by Donnan dialysis. Herein, the removal of boron by Donnan dialysis was investigated using RSM approach. RSM is a useful method for process optimization when a number of independent factors and their interactions have an impact on the desired results. With RSM, multiple variables are tested concurrently with the least amount of trials possible in accordance with unique experimental designs built on factorial designs. This technology has the advantage over conventional approaches in that it requires less time and money. The goal of RSM is to characterize the behavior of a dataset in order to make statistical predictions. It is a set of statistical and mathematical tools enabling the adjustment of experimental data to a theoretical model expressed by a polynomial equation. The simultaneous variation of a large number of operating parameters at once, the reduced number of experiments, the detection of interactions between factors, and achieving the highest precision are all advantages of the RSM for optimization [22]. Recently, many statistical experimental design methods have been employed in optimization. Among them, Doehlert designs stand out, compared to other designs such as the central composite design or Box-Behnken design, by the reduced number of experiments and the possibility to assign a large or a small number of levels to the chosen variable. Indeed, this design presents variables with different numbers of levels. The variable with the stronger effect should be the variable with the highest levels if you want to learn as much as you can about the system.

Doehlert designs are also more efficient in mapping space: adjoining hexagons can fill a space completely and efficiently since the hexagons fill space without overlap. Moreover, Doehlert design is distinguished by a low ratio between the number of coefficients and the number of experiments. Thus, it can be considered as more efficient than the central composite design or Box-Behnken design [22]. Investigations of boron removal by Donnan dialysis were carried out considering four operating factors, i.e. the counterion concentration in the receiver compartment, the boron concentration, the pH of the feed compartment, and the type of the anionic-exchange membrane. The effect of these factors and their interactions were evaluated using the full factorial design. The Donnan dialysis was then optimized using the Response Surface Methodology based on the Doehlert design.

#### **2. Methods and materials**

#### **2.1 Membranes**

In the Donnan dialysis process, two membranes have been used, which are Neosepta® AFN and Neosepta® ACS (supplied by Alstom). Before any measurement, it is necessary to condition the samples to stabilize their physicochemical properties and to eliminate any impurities that could come from their manufacturing process. **Table 1** shows the properties of the used membranes determined according to the standard NF X 45-200 [23]. The water content was determined by the Mettler-Toledo moisture thermo balance device. The water content was calculated by Eq. (1):

$$\text{WC}(\%) = \frac{\text{W}\_{\text{h}} - \text{W}\_{\text{d}}}{\text{W}\_{\text{h}}} \times 100 \tag{1}$$

where Wh is the mass of the hydrated membrane, Wd is the mass of the dried membrane, and WC(%) is the water content percentage. The water content is the difference in mass between the hydrated membrane (which has been immersed in the proper stabilization content and slightly compressed to remove the surplus liquid) and the dried membrane (which has been dried at 140°C until membrane mass stabilization indicates that all of the water has been removed).The mean value of 10 measurements at different locations using a 1�μm resolution Käfer Thickness Dial Gauge is the dry-state membrane thickness.

The ion-exchange capacity (in meq. of functional sites per gram of dry membrane or per cm<sup>3</sup> of wet membrane) was determined following the French standard NF X 45-200 [23].

In order to prepare the samples for the Donnan dialysis operations, the samples had to be conditioned before any measurement could be made. This was done primarily to eliminate contaminants from the production process and to stabilize their physical-chemical properties. French standard NF X 45-200 was followed in the conditioning process.


**Table 1.**

*Properties of the two membranes used.*

#### **2.2 Donnan dialysis (DD)**

The Donnan dialysis is a method of membrane separation in which identically charged ions are exchanged between two solutions via an ion-exchange membrane [24–26]. The chemical potential gradient acts as the driving force in Donnan dialysis; anions are exchanged stoichiometrically through an anionicexchange membrane, and the procedure is only complete if the Donnan equilibrium is attained. Since the electroneutrality is maintained, the feed should exchange the same number of anions with the receiver compartment in the opposite direction [19, 27, 28].

**Figure 1** indicates the schematic flow of Donnan dialysis. The apparatus is used to study Donnan dialysis's removal of boron. It consists of a cell with feed and receiver compartments divided by an anion-exchange membrane inside a thermoregulated water bath. A peristaltic pump with two identical heads and a speed variator that allows for varied flow rates is used to pump the solutions through the cell. Through the use of two stirring rods with variable speeds, the hydrodynamic conditions on either side of the membrane can be changed. Two removable sections constructed of polymethylmetacrylate (plexiglass) make up the dialysis cell. It consists of four pieces that are connected by three threaded rods made of stainless steel. Supports provide the centering. The two tubes that make up the two compartments at the center are symmetrical. Each compartment has three threaded holes that provide support for inserting boxes. The membrane forms a seal by sandwiching itself between these two compartments. The feed and the receiver compartment were supplied, through a peristaltic pump, with NaCl and a containing boron solution, respectively. The used membranes were AFN and ACS. Boron concentrations were determined after 7 hours of treatment by Donnan dialysis for each experiment. By reacting the samples with azomethine-H and then measuring the absorbance at 420 nm with a UV-visible spectrophotometer, the samples' boron concentration was determined [29]. A linear concentration between 1 and 4 mg/L was found. Higher concentration samples were diluted to fit the previously mentioned linearity range.

Boron removal rate was determined by Eq. (2):

$$\mathbf{Y}(\emptyset) = \frac{\mathbf{C}\_0 - \mathbf{C}\_e}{\mathbf{C}\_0} \times \mathbf{100} \tag{2}$$

With C0 and Ce are the initial boron and equilibrium concentrations, respectively.

**Figure 1.** *Schematic flow of Donnan dialysis.*

#### **2.3 Optimization process**

To identify the most significant and influencing parameter, a full factorial design was performed first, followed by an RSM design based on the Doehlert matrix. NemrodW® was the program employed in this study. The practical use of the Experimental Research Methodology (experimental designs) depends on the NemrodW® program.

#### **3. The preliminary study**

The definition of each factor's levels must be done with great care since if the domain is either too tiny or too large, the mathematical models might no longer work. Because of this, it is advised to do a preliminary study to help determine the appropriate high and low levels for each element. To establish the bounds of each parameter and better pinpoint the optimal value, a preliminary study was carried out as a phase in the parameters' pre-optimization process.

#### **3.1 The pH effect in the feed compartment**

The pH effect of the feed compartment is studied by varying the initial pH of the feed solution from 9.5 to 12.5. **Figure 2** shows the Boron removal rate under different initial pH values for a counterion concentration of 0.1 mol/L, an initial boron concentration of 50 mg/L, and a stirring speed of 500 rpm.

Due to its effect on the transfer of boron from the feed compartment to the receiver compartment, the effect of pH was first investigated. According to **Figure 2**, at a pH of 11.5 the AFN membrane removed the most boron (45%), while the ACS membrane removed 17%. This can be explained by the two species of boron that occur in aqueous solutions at various pH levels, which are the boric acid B(OH)3 in diluted aqueous solutions below pH 7 and the metaborate anion B OH ð Þ� <sup>4</sup> at pH 10 [30, 31]. However, above a pH of 11.5, the presence and competition with OH are likely to have

**Figure 2.** *Influence of pH on the removal rate of boron.*

an impact on the transport of boron, which reduces boron removal because the hydroxyl ion transport is preferred because OH has much higher mobility than boron. In reality, the boron transfer process involves three steps. First, the boron in the feed solution is exchanged with ions or ionizable groups of the anion-exchange membrane. The second step involves the transport of boron across the membrane to the receiving solution side. In the third step, the boron is transported into the receiver solution following an exchange with the counterions to guarantee electroneutrality [32, 33]. Therefore, it can be said that for the two used membranes, the highest boron transport was attained at pH = 11.5 in this case.

#### **3.2 Concentration of chloride in the receiver compartment effects performance**

One of the factors influencing the elimination of boron through the anionicexchange membrane during the Donnan dialysis procedure is the chloride concentration. To study the effect of this parameter, the concentration of the Cl counterion was varied from 0.01 mol/L to 0.05 mol/L in the receiver compartment for a boron concentration fixed at 50 mg/L. **Figure 3** displays the impacts of the two membranes' receiver compartment's chloride concentration.

According to **Figure 3**, the flux of boron ions through anion-exchange membranes increases with the increase of chloride concentration from 0.01 to 0.5 mol/L. At 0.01 mol/L of counterion, the boron removal rate reaches only 32% and 13% for AFN and ACS, respectively. At 0.5 mol/L of the concentration of Cl, the removal efficiency increases to 58% for AFN and 38% for ACS. The improvement in the cross-ion transfer between Cl and boron necessary to maintain electroneutrality is explained by the fact that the concentration gradient of the counterions grows.

For the two membranes, it appears that the improvement in the boron removal in the feed compartment, as shown by an improvement in the exchange's kinetics, is related to the increase in the concentration of counterions in the receiver compartment. In fact, it is known that ion exchange is faster when the concentration of counterions is higher in the receiver compartment [34–36].

**Figure 3.** *Concentration of chloride in the receiver compartment effects performance.*

#### **3.3 Boron concentration effect**

The elimination by Donnan dialysis is significantly influenced by the amount of boron in the feed compartment. Under these circumstances, the receiver compartment's Cl- concentration is 0.5 mol/L, the feed compartment's pH is 11.5, and the boron concentration rises from 5 mg/L to 100 mg/L. **Figure 4** presents the findings.

According to **Figure 4**, the improvement in cross-ion transfer between Cl� and B OH ð Þ� <sup>4</sup> helps to maintain the gradient concentration of boron at a high level. The elimination was 40% for AFN and 17% for ACS at the feed compartment's lowest boron content (25 mg/L). When the boron content is raised to 100 mg/L, their elimination is improved to 75% with AFN and to 48% with ACS. A similar result was reported by Tor [37].

#### **3.4 Membranes choice**

According to the results of **Figures 3** and **4**, the boron removal rate by Donnan dialysis depends significantly on the AEM properties. Therefore, the most effective membrane is AFN since it allows to reach 75% after 7 hours of treatment against 48% for ACS membrane. According to Akretche, (i) a high exchange capacity boosts the selectivity between monovalent and multivalent anions due to the higher repulsion charge, (ii) a high thickness reduces diffusion, resulting in a lower ions flux, and (iii) a high water content can reduce permselectivity and encourage the penetration of bulky ions [38]. In actuality, the AFN membrane exhibits the largest water content, the highest ion-exchange capacity, and a higher permeability to monovalent than bivalent anions. On the other hand, the ACS has the lowest permeability due to its large thickness and low water content. The works of Ayyildiz [31], who reported that the removal of boron by Donnan dialysis is more successful using the AFN membrane, support this conclusion. AFN membrane has been chosen as the subject of the following investigation.

**Figure 4.** *Boron concentration effect.*

#### **4. Full factorial design**

To ascertain the impact of these variables and how they interact with one another on the removal of boron by Donnan dialysis, the full factorial design was used. We were able to establish the experimental field and the level that had to account for every element thanks to the preliminary study. The starting boron concentration, counterion concentration, and feed compartment pH were the three variables that were selected. In order to more clearly define the examined response (boron removal efficiency), restrictions are imposed. The membrane AFN was used in the Donnan dialysis procedure.

A full factorial design was used to assess how operating parameters affected the removal of boron by Donnan dialysis. **Table 2** displays the experimental ranges and factors level. For each of the three component designs specified in the study, a full factorial matrix made up of eight distinct experiments was used. A linear polynomial model with interaction is used to model the experimental response related to a factorial design (see Eq. (3)):

$$\mathbf{Y} = \mathbf{b}\_0 + \mathbf{b}\_1 \mathbf{X}\_1 + \mathbf{b}\_2 \mathbf{X}\_2 + \mathbf{b}\_3 \mathbf{X}\_3 + \mathbf{b}\_{12} \mathbf{X}\_1 \mathbf{X}\_2 + \mathbf{b}\_{13} \mathbf{X}\_1 \mathbf{X}\_3 + \mathbf{b}\_{23} \mathbf{X}\_2 \mathbf{X}\_3 \tag{3}$$

where Y is the experimental response, Xi denotes the coded variable, bi denotes the estimation of factor i's principal effect on the response Y, and bij denotes the estimation of factor i and factor j's interaction effect on the response Y.

The coefficients of the model were estimated in accordance with the findings in **Table 3**, and it was discovered that (see Eq. (4)):


**Table 2.**

*Experimental range and factors level studied in the factorial design.*


**Table 3.** *Full factorial design matrix.* *Boron Removal by Donnan Dialysis According Doehlert Experimental Design DOI: http://dx.doi.org/10.5772/intechopen.111869*

**Figure 5.** *Pareto analysis of the removal of boron*.

$$\mathbf{Y(\%)} = \mathbf{34.85} + 2.50 \,\mathbf{X\_1} + \mathbf{3.75} \,\mathbf{X\_2} + 6.15 \,\mathbf{X\_3} - 0.20 \,\mathbf{X\_1} \mathbf{X\_2} - 0.90 \,\mathbf{X\_1} \mathbf{X\_3} - \mathbf{1.15} \,\mathbf{X\_2} \mathbf{X\_3} \tag{4}$$

The different coefficients of the polynomial model (Eq. (4); R<sup>2</sup> = 0.999) were determined, which represented the effects and interactions of the various investigated factors. The Pareto analysis (**Figure 5**) allows to evaluate the contribution of each parameter on the response according to the equation (Eq. (5)):

$$\mathbf{P\_i} = \left(\frac{\mathbf{b\_i^2}}{\sum \mathbf{b\_i^2}}\right)^2 \times 100\tag{5}$$

The three investigated factors have a favorable impact on the observed behavior, i.e., increasing them improved boron removal. In contrast to the 17.9% they contributed to pH, and their contributions to the examined response were just 6.6% for boron concentration and 2.9% for chloride concentration. Thus, two factors—pH and boron concentration—can have a significant impact on the elimination. The solution pH coefficient's positive value indicates that boron removal was enhanced. This is brought on by the existence of, which, high pH levels, becomes the dominant species. The elimination of boron by Donnan dialysis is moderately affected by both the counterion concentration and the boron concentration. Therefore, the feed compartment's pH is the most crucial variable.

#### **5. Doehlert design**

In this investigation, the optimum condition was found using the Response Surface Methodology (RSM) in accordance with the Doehlert design. By evenly dispersing the experimental points within the space-filling of the variables, Doehlert's method is created. N = k<sup>2</sup> + k + 1 is the total number of experiments for k factors. Fifteen tests were in total, three of which were replicated in the central field [39, 40]. The initial boron concentration, feed compartment pH, and receiver compartment counterion


#### **Table 4.**

*Experimental range and levels of the factors.*

concentration were all factors that were examined. According to the preliminary study, these parameters' upper and lower limits were established. To get the most information out of the system, it is typically preferable to use the variable with the significant effect as the variable with the highest levels. The experimental field of the factors under investigation is shown in **Table 4**.

An experiment conducted in the Doehlert domain is able to predict, at any point in the experimental domain, the value of an answer by estimating the coefficients of a second-order function [40]. The selected model uses a polynomial equation (see Eq. (6)) to describe the predicted values of the responses Y:

$$\mathbf{Y} = \mathbf{b}\_0 + \mathbf{b}\_1 \mathbf{X}\_1 + \mathbf{b}\_2 \mathbf{X}\_2 + \mathbf{b}\_3 \mathbf{X}\_3 + \mathbf{b}\_{11} \mathbf{X}\_1^2 + \mathbf{b}\_{22} \mathbf{X}\_2^2 + \mathbf{b}\_{33} \mathbf{X}\_3^2 + \mathbf{b}\_{12} \mathbf{X}\_1 \mathbf{X}\_2 + \mathbf{b}\_{13} \mathbf{X}\_1 \mathbf{X}\_3 + \mathbf{b}\_{23} \mathbf{X}\_2 \mathbf{X}\_3 \tag{6}$$

The estimated principal effect of factor i is denoted by the letters bi, the estimated second-order effects by the letters bii, the estimated interactions between factors i and j by the letters bij, and the coded variable by the letters Xi.

The regression coefficients (R<sup>2</sup> ) and the percentage absolute errors of deviations (AED) between experimental and calculated results must be considered for model validation. The AED was calculated from Eq. (7):

$$\text{AED} \left( \% \right) = \frac{\mathbf{100}}{\mathbf{N}} \cdot \left| \frac{\mathbf{Y\_{exp}} - \mathbf{Y\_{theo}}}{\mathbf{Y\_{exp}}} \right| \tag{7}$$

where Yexp and Ytheo are the responses obtained from experiments and from the model, respectively. N is the number of points at which measurements were carried out. A model was considered valid if R2 > 0.7 and AED < 10% [41].

For boron removal optimization, Response Surface Methodology via Doehlert design was used with membrane AFN. To optimize the factors, 15 experiments involving three variables were evaluated using a Doehlert experimental design (**Table 5**).


*Boron Removal by Donnan Dialysis According Doehlert Experimental Design DOI: http://dx.doi.org/10.5772/intechopen.111869*


#### **Table 5.**

*Doehlert Matrix and obtained results.*

Using the experimental results from **Table 5**, the second-order polynomial equation was fitted to the data appropriately and the coefficients were presented in Eq. (8):

$$\mathbf{Y} = \mathbf{84.2} + \mathbf{3.01X\_1} + \mathbf{18.81X\_2} + \mathbf{9.12X\_3} + \mathbf{0.20X\_1^2} - \mathbf{65.90X\_2^2} - \mathbf{37.40X\_3^2} \tag{8}$$

$$- \mathbf{2.38b\_{12}X\_1X\_2} - \mathbf{010} \, \mathbf{X\_1X\_3} - \mathbf{12.20X\_2X\_3}$$

Based on the obtained results, the coefficients show that pH of the feed compartment had a significant effect on boron removal (b2 = 18.81). As a second influencing factor, boron concentration (b3 = 9.12) was taken into consideration. However, chloride concentration had a less significant effect on boron removal (b1 = 3.11). The feed compartment's pH and boron concentration (b23 = �12.20) had the most significant interaction and had an adverse impact on Donnan dialysis's ability to remove boron.

Although they had a small impact on the removal of boron by Donnan dialysis, the interactions between the chloride concentration and the feed compartment's pH (b12 = �2.38) and the concentration and boron concentration (b13 = �0.10) were not.

The regression coefficient (R2 ) and the percentage of absolute errors of deviation (AED) were used to evaluate the model's validity. The regression coefficient (R2 ) is better than 0.7, and the percentage of absolute errors of deviation (AED) (%) = 0.425% was less than 10%.

The contour plots (curve of constant response) are used to describe how boron is removed by Donnan dialysis. The response surface is represented in a contour plot as a two-dimensional plane where all points with the same response are joined to form contour lines with constant responses. Typically, a surface plot shows a threedimensional image that could give a clearer idea of the response. To show the relationship between two factors and a response, use contour plots. The graph shows values of the pH for combinations of the [Cl�] and B OH ð Þ� <sup>4</sup> . The [Cl�] and B OH ð Þ� <sup>4</sup> values are displayed along the X- and Y-axes, while contour lines and bands represent the response value Y. The obtained plots are illustrated in **Figure 6**.

At a constant boron content of 62 mg/L, the first plot displays the combined fluctuation of chloride concentration and pH. The contour plots' form demonstrates that, only at pH 11.5 when chloride concentration increases from 0.1 mg/L to 0.3 mg/L,

**Figure 6.** *Contour plots and three dimensions plots.*

boron removal improves. This was attributable to the many boron forms that can exist in aqueous solutions at various pH levels. At higher pH levels, the B OH ð Þ� <sup>4</sup> is the dominating species. However, beyond pH 11.5, the presence and competition with OH- , which reduces the removal of boron, are likely to have an impact on the transport of boron. Therefore, it can be said that pH 11.5 produced the largest boron transfer.

The variation in boron concentration and pH at a fixed chloride concentration of 0.3 mol/L is shown in the second plot. The shape of these iso-response curves, which are concentrated in the center of the domain, demonstrates that the pH has a significant impact on the elimination of boron. This was anticipated because boron removal was positively influenced by the pH and chloride content. The variation of chloride and boron concentrations with a constant pH of 11.5 is depicted in the third plot. According to the contour plots, the rate of boron removal improves only when the chloride concentration is increased from 0.1 mg/L to 0.3 mg/L around the boron concentration of 62 mg/L.

The optimum values are 66 mg/L for the concentration of boron, 0.5 mol/L for the concentration of chloride, and 11.6 for the pH of the feed compartment. These conditions led to a maximum removal of boron of 88.8%. A replicate three times of experiment was conducted in the optimum conditions in order to verify the efficiency of predicting values. The coefficient of repeatability is less than 1%, so it can be concluded that the removal of boron by Donnan dialysis is reproducible.

#### **6. Conclusion**

The influence of operating parameters on boron removal by Donnan dialysis was investigated using two different membranes, AFN and ACS. For an initial boron

*Boron Removal by Donnan Dialysis According Doehlert Experimental Design DOI: http://dx.doi.org/10.5772/intechopen.111869*

concentration of 100 mg/L, a pH of 11.5 in the feed compartment and 0.5 mol/L of Cl-, the counterion, in the receiver compartment, 75% of boron removal rates for AFN and 48% for ACS were recorded. The influence and interactions of these parameters were then evaluated using a full factorial design. It was concluded that pH is the most influent in the elimination of boron. The Response Surface Methodology by Doehlert enabled the identification of the ideal working conditions for the removal of boron, which reached an efficiency of 88.8% using an AFN membrane, which were [B] = 66 mg/L, pH = 11.6 and [Cl-] = 0.5 mol/L. Compared to the conventional "oneat-a-time" approach, using the Response Surface Methodology to identify the optimal conditions for 13.8% can be seen of as a good option.

#### **Acknowledgements**

The researchers would like to thank the ICMPE/University Paris Créteil Est for funding the publication of this project.

### **Author details**

Ikhlass Marzouk Trifi<sup>1</sup> \*, Lasâad Dammak<sup>2</sup> , Lassaad Baklouti<sup>3</sup> and Béchir Hamrouni<sup>1</sup>

1 Desalination and Water Treatment Research Laboratory, Faculty of Sciences of Tunis, University of Tunis El Manar, Tunis, Tunisia

2 Institute of Chemistry and Materials Paris-Est (ICMPE), Paris-Est University, Thiais, France

3 Department of Chemistry, College of Sciences and Arts at Al Rass, Qassim University, Saudi Arabia

\*Address all correspondence to: ikhlassmarzouk@gmail.com

© 2023 The Author(s). Licensee IntechOpen. This chapter is distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/3.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.

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#### **Chapter 3**

## Boron Nutrition in Horticultural Crops: Constraint Diagnosis and Their Management

*Pauline Alila*

#### **Abstract**

Out of 30 elements, 16 horticulture crops require them to thrive. All horticultural crops require boron, one of the necessary elements, to function. Extremely trace amounts of boron are present in soils. The majority of the boron that is readily available in humid areas is primarily contained in organic debris, which is broken down by microorganisms for the benefit of plants. In the tropics this element is leached down in soil due to heavy rainfall. As trace element B plays an important role in the growth and development of plants. Various crops exhibit symptoms of deficiency as well as of toxicity when there is even a slight aberration of available boron in soil. Therefore, it is imperative to study and understand the optimum requirement of B by specific crops. Boron also interacts with other elements and manifests in crop plants in various ways. This chapter attempts to understand some of the roles of boron in horticultural crops (fruits and vegetables) and its management for optimum growth and development in crop plants.

**Keywords:** boron, its importance, diagnosis, horticultural crops, management

#### **1. Introduction**

Among the essential elements, boron plays an important role in all the horticultural crops. The microbial decomposition of organic matter in soil releases the available boron for use in plants particularly in humid region. Most of the different macro and micronutrients become exhausted in soil due to continuous uptake of these elements by the crops, thereby causing deficiency in plants which manifests in nutritional disorders often resulting in low yields. Warington in the year 1923 [1] demonstrated the essentiality of boron (B) for the growth and development of higher plants. Boron is crucial as a vital micronutrient for achieving optimal crop growth, development, yield, and quality. It plays a key role in forming and stabilizing cell walls, promoting lignification, and facilitating xylem differentiation [2]. Moreover, boron is essential for enhancing the protein and enzymatic functions of cell membranes, thereby improving membrane integrity. It's worth noting that sexual reproduction in plants is more vulnerable to boron deficiency compared to vegetative growth. During the pollination of flowers, boron supports pollen tube growth, ensuring successful seed set and fruit development. Additionally, boron contributes to increased nectar

production in flowers, attracting pollinating insects. In the realm of crop production, boron stands out as a critical micronutrient required for the healthy growth of most crops. Furthermore, boron imparts drought tolerance to plants. Among the essential micronutrients, boron is unique as the sole non-metal present in a non-ionic form. Commercially, boron is sourced from minerals such as ulexite, natural boric acid (sassolite), borax (tincal), colemanite, and kernite, with the United States (US) and Turkey serving as the primary and richest sources of this element.

In vascular plants, known as tracheophytes, boron follows a path from the roots through the transpiration stream to the points of active growth, where it collects in the stems and leaves. It has been proposed that the localized accumulations of boron in these growing tissues have contributed to the evolutionary development of a reliance on boron for certain metabolic processes in plant meristems. Once boron accumulates in leaves, its subsequent movement within the plant is limited, and it becomes fixed in the apoplast. What sets boron apart from other nutrients is the considerable variation in its mobility among different plant species. The mobility of boron is facilitated by its interaction with polyols. In species where polyols (such as sorbitol, mannitol, and dulcitol, which are water-soluble sugar alcohols) are the primary products of photosynthesis, boron can move rapidly and effectively through the phloem. Examples of such species include those from the Malus, Prunus, and Pyrus genera (including apple, prune, plum, peach, pear, cherry, almond, and apricot), as well as olive and coffee in fruit plants, and carrot, onion, pea, celery, bean, cauliflower, cabbage, asparagus, wheat, and barley in vegetables and cereals. Typically, boron concentrations are highest at the tips and edges of leaves, but in species where boron is phloem-mobile, the concentration remains consistent throughout the leaves. Once again, it is suggested that boron's movement through the phloem is limited when boron levels are higher in older leaves. The symptoms of boron toxicity, which manifest at the edges of older leaves, have been interpreted as evidence of boron's immobility within plants [3]. Conversely, when there is a higher concentration of boron in young leaves and fruits, it serves as an indicator of phloem mobility [4–7]. Research indicates that maintaining optimal levels of boron enhances the germination of pollen and the growth of pollen tubes in almond trees, leading to improved fruit development and seed growth [8]. Applying boron as a foliar treatment to sour cherry trees just before the leaves fall significantly boosts boron levels in flower buds and enhances fruit set [9]. When boron is applied to trees in the autumn, it translocates from the leaves to the adjacent buds, where elevated levels persist and become evident in the flowers during anthesis. Generally, flowers receive their boron supply from reserves in the wood, which become mobilized during the development of the flowers. Boric acid, being a soluble compound susceptible to leaching by rainfall, frequently results in boron deficiency in regions with high rainfall and humidity. In contrast, soil boron toxicity is less common and tends to occur in arid and semi-arid areas. The concentration and availability of boron in soils are influenced by various factors, including the soil's parent material, texture, clay mineral composition, pH levels, liming, organic matter content, and its interactions with other elements. Consequently, understanding these factors that affect boron uptake is essential for assessing boron deficiency and toxicity under different environmental conditions.

#### **2. Diagnosis of boron deficiency by soil and plant analysis**

As early as 1942, Dregne and Power [10], reported that B availability in the soil is influenced by numerous factors such as soil reaction, soil moisture, active calcium and

#### *Boron Nutrition in Horticultural Crops: Constraint Diagnosis and Their Management DOI: http://dx.doi.org/10.5772/intechopen.113367*

organic matter content. Boron deficiency in soil is mostly found in acidic soils, irrespective of parent material where excess B is leached out. Soils arising from igneous and metamorphic sandstone rocks are found to be naturally low in B. soils deficient in B are also found in laterite soils which have limited silica and strong Fe and Mn. Although alkaline soils are rich in total B but poor in available B for plants. Similarly, in peat soils and acid sludge and soil with low clay composition B deficiency is evident. In general, it is estimated that the total B concentration is between 20 and 200 mg B/kg, where its available concentrations vary greatly in different soil types. Boron is available to plants in water soluble form in soil. It is naturally present in the soil in the water-soluble form, which is readily available as a nutrient for plants. Movement of boron in soil is through "mass flow" which is absorbed by plants through transpiration stream as in water absorption. Consequently, in dry soils, boron uptake may be reduced or unavailable at a time of maximum B requirements in plants. Certain environmental conditions can also lead to low availability of soil boron. High rainfall can cause leaching of available boron from the root zone which is a major problem, especially in coarse-textured soils and when plants are at or just before the rapid growth of leaves and flower development stages. The other adverse environmental condition is when less rainfall or drought period occurs just prior to, or during flowering and seed set.

The requirements for boron (B) in plant growth exhibit significant variability across different plant species, as well as within the same species at various growth stages [2, 11–14]. In soil, a boron level below 0.5 mg/kg of water-soluble boron is considered low or deficient. An optimal or medium range of boron falls within the range of 0.5–2 mg/kg, while soils containing more than 2 mg/kg are deemed to have a high or excessive boron content. Consequently, soils containing less than 0.5 mg/ kg of hot water-soluble boron are generally incapable of providing sufficient boron to support normal growth and yield in most plant species.

The total boron content in Indian soils has been observed to vary between 7 and 630 mg B/kg [15]. Reports of boron deficiency have primarily emerged from the soils in Indian states such as Assam, Bihar, Meghalaya, West Bengal, Jharkhand, and Odisha. This deficiency is notably prevalent in acid-red and lateritic soils, including high-pH calcareous soils [16]. Light-textured soils, like sandy loam and loamy sands, also tend to lack sufficient boron content due to their excellent drainage properties, which result in effective leaching [17]. Given that boron is a weakly held anion, it can be easily washed out of the soil, rendering acid-sand soils particularly susceptible to boron deficiency [18].

Leaf analysis, supplemented by soil analysis, can be reliable diagnostic tool for analysis of B availability and status in plants. The leaf analysis is derived from four factors:


Boron, which plants absorb in the form of undissociated boric acid (B (OH)3 or H3BO3), possesses a strong capacity to create complexes within the plant system.

In the case of most crops, a boron content ranging from 15 to 100 mg/kg in plant tissue is considered sufficient for regular growth. Conversely, levels exceeding 200 mg/ kg may be excessive, leading to a potentially toxic or inhibitory impact on crop growth and yields. When the concentration of boron in plant tissue falls below a critical threshold, it hinders the optimal growth and yield of the crop. Plants have quite specific demands for boron, and the gap between excess and deficiency is quite narrow. For instance, maintaining a leaf boron concentration within the range of 30–70 μg/g is ideal in mini cucumbers. Deficiency becomes evident when levels drop below 20 μg/g, while toxicity is observed when concentrations exceed 100 μg/g. Notably, boron does not readily migrate from older to newer plant tissues, making the roots to continuously uptake essential for the plant to grow normally. The importance of boron in pollination and in regulation of cells development is well established and its deficiency leads to poor seed set and fruit development. The cost of boron in correcting boron deficiency is justified with improvement in both the growth and yields of crop [19].

#### **3. Interaction of boron with other elements**

There has been reports of interactive effect of boron in plants with the other nutrient elements. In general, excessive applications of potassic fertilizers results in high potassium concentration which induces boron deficiency. B has been found to counter the toxic effects of aluminum on root growth of dicotyledonous plants [20]. In limestone soils rich in soil calcium reduces the availability of boron. Conversely, when there are excessive boron levels in soil, plant toxicity due to boron can be prevented by calcium application. Reports on the application of zinc to neutralize toxic effects of boron in some plants and there was subsequent increase in the crop yield is documented [19].

#### **4. Correction of boron deficiency**

When plants display deficiency symptoms of Boron, the issue can be easily addressed by administering borax, which is a white crystalline salt that is needed in very small amounts. Applying 5–10 kg/ha of boron can rectify boron deficiency in most plants. Boron can be delivered through both foliar and soil fertilization methods to ensure an adequate supply to plants. Foliar application is most effective when root activity is restricted and boron deficiency symptoms become evident during dry soil conditions in the growing season. Generally, a 0.2% borax solution can be sprayed onto the foliage every 10 days until the deficiency is corrected. For many vegetable crops, Solubor (100 g/100 L of water) can also be utilized as a foliar spray [19]. However, Das et al. [21] noted that soil application of boron outperforms foliar sprays. In calcareous soils, introducing sulfur at a rate of 2 MT/ha helps reduce soil pH to the range of 6.0–7.0 and enhances the boron solubility in the soil solution. To address hidden deficiencies, a spray of 0.2% boric acid or borax during the pre-flowering or flower head formation stages has been shown to increase yields. Typically, the application of boron ranges from 0.25 to 3.0 kg/ha, depending on the specific crop requirements and the chosen application method. Broadcast applications typically necessitate higher rates compared to banded soil applications or foliar sprays. It's crucial to exercise caution when addressing boron-deficient crops, as boron is highly toxic to most plants at relatively low levels. Additionally, ensuring an

even distribution across the soil is essential to prevent crop plants from experiencing toxicity. In general, applying 10 kg of borax (Granubor) per hectare to deficient soil before planting is an effective preventive measure against boron deficiency.

#### **5. Effect of boron in vegetable crops**

Boron requirements vary among different plant species, and this variability may be attributed to differences in cell wall composition. Cereals and grasses exhibit lower sensitivity to low levels of available boron compared to legumes and certain vegetable crops. In dicots such as soybean and alfalfa, it's observed that the deficiency concentration of boron is three to four times higher in younger leaves than in older ones, which indicates the limited mobility of boron through the phloem in these species. Given boron's immobility within plants, soils with marginal boron levels can lead to crop deficiencies during critical growth phases, namely, during vegetative growth, flowering, and seed development. Consequently, maintaining a consistent boron supply during the growth season is crucial for achieving optimal growth and the seed yield. Experimental evidence demonstrates that boron significantly enhances yield, mobility, and stress tolerance in various crop species (see **Table 1**). In extreme cases, crops grown in low-boron soils perform well until the flowering stage, at which point there can be substantial yield losses due to floral abortion or failed seed set. The most effective time for boron application is at the beginning of the reproductive phase, considering its immobility within plants, and this timing depends on the photosynthetic efficiency of the specific plants. Some examples of deficiency symptoms are outlined in **Table 2**.

The issue of hollow stem in cauliflower was effectively addressed by applying Farm Yard Manure at a rate of 7.5 tons/ha as a basal application, coupled with either the use of Boric Acid at 0.3% or Liquid Boron at 1.5 g/l, carried out 30 days after planting. Notably, the application of Liquid Boron was found to be the more costeffective choice. This boron application significantly resolved these problems and


#### **Table 1.**

*Influence of B application on various crop plants [22].*


#### **Table 2.**

*Visible symptoms of boron deficiency in some vegetable crops.*

led to an increase in curd yield [34]. In the case of muskmelons, applying boron improved both plant growth and fruit quality. Additionally, it reduced the occurrence of chilling injury in harvested fruit during cold storage at 5°C [33]. An optimal treatment combination consisting of 120 kg N/ha along with 0.6 kg B/ha was identified for achieving the maximum yield of tomatoes [35]. Furthermore, Naz et al. [36] reported that applying 2 kg B/ha resulted in the highest number of flower clusters per plant, fruit set percentage, total yield, fruit weight retention, and total soluble solids for the Rio Grand and Rio Fique tomato cultivars.

In a four-year study assessing the direct and residual effects of applied boron (B) in French bean-cauliflower cropping sequences, it was observed that applying 8 kg/ ha of B led to reduced crop yields in eight crops spanning 4 years. When 2 kg/ha of B was applied, it enhanced French bean yields in soils with low-available B, whereas high-available B soils initially reduced yields in the first two crops but improved yields in the third and fourth crops. However, in both French beans and cabbage, elevated levels of B in plant tissue resulted in toxicity. The hot water-soluble B (HWS-B) content at harvest for each crop indicated a rapid decrease in the availability of applied B in these soils. Applying high levels of B-containing fertilizer to these soils led to the accumulation of B in toxic concentrations, resulting in lower vegetable yields.

#### *Boron Nutrition in Horticultural Crops: Constraint Diagnosis and Their Management DOI: http://dx.doi.org/10.5772/intechopen.113367*

For French beans, which are low accumulators of B and more sensitive crops, it is advisable to rely on residual B rather than applying B-containing fertilizer directly to the soil in any vegetable cropping system, especially in red soils [37].

Solanki et al. [38] found that the application of boron at rates of up to 1 kg B/ha led to a significant increase in the yields and dry matter production of vegetable crops, including carrot, cauliflower, and onion. However, when the boron level was raised to 2 kg B/ha, yields tended to decline. The extent of this response varied from one crop to another and followed a descending order of magnitude: carrot > cauliflower > onion. Additionally, the application of boron resulted in an improvement in both the content and yield of protein in vegetable crops compared to the control. Over time, the introduction of boron into the soil led to a progressive increase in its concentration and uptake by vegetable crops. The highest removal of boron was observed in cauliflower curds, while the lowest was noted in carrot roots. The percentage of apparent boron recovery was influenced by its levels, with the highest recovery occurring at the 1 kg B/ha level. However, as boron levels increased, boron use efficiency decreased, with the lowest efficiency recorded at 1.5 kg B/ha.

#### **6. Effect of boron on fruit crops**

Boron has been identified as a crucial element influencing reproductive processes, impacting anther development, pollen germination, and pollen tube growth. Apple trees, scientifically known as *Malus domestica* Borkh., are particularly recognized for their high demand for boron [14]. The initial visual indicators of boron deficiency are evident in poor fruit set, which subsequently leads to reduced yields. This deficiency is critical because boron plays a pivotal role in reproductive growth [39]. In apple trees suffering from boron deficiency, the fruits tend to be undersized, corky, distorted, and prone to cracking and russeting. They exhibit yellow skin and fail to develop a healthy red coloration [40]. Additionally, apple fruits may have diminished concentrations of soluble solids and acids when boron levels are insufficient [41]. Stone fruits are likewise affected by boron deficiency. For instance, cherry shoots that lack adequate boron growth initially but subsequently undergo necrosis at the tips and ultimately die. In B-deficient plants, some buds may remain closed during springtime, while others wither and perish. The presence of cracking, deformation, shriveling, both internal and external browning, as well as corking around the pit and in the flesh, serves as clear indicators of boron deficiency in cherry fruit [42]. Similar responses to low-boron soils are also observed in nut crops grown in temperate regions.

Boron stands as an indispensable mineral for the growth and proper functioning of citrus plants; however, it is frequently deficient in many types of soil. Pronounced deficiency, ranging from 39 to 68%, is notably prevalent in red and lateritic soils, as well as in leached acidic soils within the hot semi-arid ecoregion. It is also observed in soils derived from alluvium within the hot sub-humid ecoregion, brown and red hill soils in the warm, humid ecoregion, and highly calcareous soils within the hot sub-humid ecoregion. Interestingly, these are the specific climate and soil combinations that yield abundant high-quality citrus crops in countries like Brazil, China, and Japan. Ruchal et al. [43] reported an increased fruit set in mandarin trees in response to higher micronutrient concentrations in their application. This could possibly be attributed to the enhanced translocation of essential nutrients and hormones to the ovary tissue, thereby stimulating fruit formation. Another potential factor

contributing to this effect could be the improved availability of microelements, which enhances photosynthesis, reduces fruit drop, and results in improvements in fruit size and quality. Assam lemon (*Citrus limon* (L.) Burm.), an indigenous lemon cultivar of Assam, is extensively grown on the warm southern slopes of the Himalayas in northeastern India. This particular lemon variety is characterized by its ability to bear fruits in multiple flushes throughout the year, necessitating adequate nutrition to achieve optimal yields with high-quality fruits. Sheikh et al. [44] noted that the treatment involving ZnSO4 (0.2%), FeSO4 (0.2%), Borax (0.2%), and CuSO4 (0.2%) led to improved lemon fruit yields and quality. This treatment also resulted in the highest number of fruits per plant (73), yield per plant (11.5 kg), fruit fresh weight (158 g), fruit length (9.60 cm), fruit diameter (5.80 cm), juice content (152 mL/fruit), TSS (6.40°B), ascorbic acid content (49.10 mg/100 g), total sugar (6.30%), reducing sugar (3.90%), non-reducing sugar (2.40%), and the lowest titratable acidity (3.13%).

Guava trees that were treated with boric acid at concentrations of 0.1% and 0.2% exhibited notable improvements in various aspects of growth and fruit development. These improvements included increased extension of the terminal shoot, greater leaf count, enhanced leaf area per shoot, and accelerated fruit ripening, with reductions of 7 and 11 days, respectively. Additionally, the yield increased by 82 and 73%, respectively, with the higher concentration producing a slightly lower yield increment. Furthermore, the fruit size saw an increase when treated with 0.1% boric acid [45]. Pre-flowering applications of boric acid at concentrations of 0.1, 0.2, 0.3, or 0.4% on Allahabad Safeda guava resulted in significant enhancements in growth, flowering, and fruiting processes [46]. For Guava cv. L-49, the largest fruits, measuring 6.68 × 7.12 cm and weighing 125.8 g, were obtained with the application of 3.0% urea and 0.3% boric acid [47]. Similarly, spraying borax at a concentration of 0.2% effectively increased the size of Sardar guava fruits, their weight (95.25 g), and the yield (63.49 kg/tree) [48]. Combining the spray of borax at 0.2% with urea at 2% in three applications (pre-flowering, fruit setting, and 3 weeks after fruit setting) for Allahabad Safeda guava resulted in fruits measuring 4.84 cm in length, 5.00 cm in width, weighing 72.67 g, and yielding 19.08 kg per tree. Alternatively, foliar applications of borax at 0.2% alone recorded a higher fruit weight of 80.67 g and a yield of 20.17 kg per plant [49]. Yadav [50] observed that the best yield of high-quality fruits (67.7 kg per tree), the highest number of fruits per tree (686), and the largest fruit volume (107.5 cc) were achieved with foliar application of a combination of urea (3.0%), borax (0.15%), and NAA (10 ppm) in guava trees. Furthermore, a preharvest spray of borax with concentrations ranging from 0.2 to 1.2% applied twice in October resulted in improved guava fruit quality in the Sardar cultivar, particularly in terms of size and weight. In another experiment involving foliar application of H3BO3 at concentrations of 0.3, 0.5, and 1.0% on guava cultivar L-49 during the winter season [51], fruit weight and yield both increased, with the highest values recorded at 1.0% B, reaching 141.0 g and 73.0 kg/tree, respectively.

The most effective method for reducing fruit drop in rose-scented litchi, as reported by Ref. [52], was the application of borax. Through foliar sprays of borax at concentrations of 0.5 and 1.0%, fruit drop was reduced to 75–76%, compared to the 92.4% fruit drop observed in the control group. In the case of litchi, Haq and Rab [53] found that foliar application of CaCl2 and borax led to significant increases in the average fruit skin calcium content (from 4.79 mg/100 g DW in the control to the highest 8.88 mg/100 g DW with CaCl2 3% + boron 1.5% treatment), boron content (from 0.109 mg/100 g DW in the control to 0.247 mg/100 g DW with the same treatment), and skin strength (from 2.43 kg/cm2 in the control to 3.01 kg/cm2 with the

#### *Boron Nutrition in Horticultural Crops: Constraint Diagnosis and Their Management DOI: http://dx.doi.org/10.5772/intechopen.113367*

same treatment). In addition, ion leakage (from 35.17% in the control to 16.17%) and fruit cracking (from 25.40% in the control to 11.14%) were reduced with the CaCl2 3% + boron 1.5% treatment. Boron plays a role in sugar movement and promotes the formation of fruit buds in plants. In the context of litchi cultivation, the use of borax at a concentration of 0.4% resulted in minimal fruit drop (69.45%), reduced fruit cracking (4.63%), and lighter seed weight (2.30 g). Furthermore, this treatment led to maximum fruit retention (30.55%), fruit set (62.50%), larger fruit dimensions (4.50 cm in length and 3.96 cm in width), greater fruit weight (24.85 g), higher pulp weight (20.73 g), an elevated fruit pulp-to-seed ratio (5.50%), increased fruit yield (120.85 kg per plant), and superior quality characteristics. These quality attributes included higher total soluble solids (22.55°Brix) and increased total sugar content (18.42%) with a lower percentage of titratable acidity (0.41%). These findings were observed in the plains of central Uttar Pradesh, India [54].

Rana and Sharma [55] discovered that grape berries and clusters exhibited an increase in both weight and volume when grapevines were subjected to boron spraying at concentrations of 0.025 and 0.05%. Furthermore, the application of calcium and boron, either individually or in combination, led to larger fruit size and a higher number of fruits per plant in the ber cv. Dongzao, consequently enhancing the fruit yield per plant [56]. In the case of Indian gooseberry and ber, the introduction of boron also contributed to an elevation in vitamin C content [57, 58]. This improvement in fruit quality may be attributed to the role of boron in facilitating carbohydrate transport within plants. Dhaker et al. [59] reported that the use of a foliar spray containing 0.6% borax significantly increased fruit weight (962.0 g) and fruit yield (21.21 kg) in bael (*Aegle marmelos* Corr.). Additionally, this concentration resulted in minimal fruit cracking (2.14%) and reduced peel thickness (2.41 mm), while also enhancing various qualitative characteristics of the fruit. Furthermore, the combined effect of organic manure (50 kg per tree) and foliar application of 0.6% borax significantly increased tree height (49.20 cm), stem girth (2.75 cm), fruit weight (980.0 g), and fruit yield (36.34 q/ha) compared to control plants.

Sotomayor et al. [60] reported that kiwifruits derived from shoots with borontreated leaves exhibited a 14.1% increase in weight compared to the control, while fruits from boron-treated flowers were 17% heavier than those from untreated flowers. Additionally, significant differences in fruit length were observed between treated and control plants. In the case of papaya cv. Shahi, foliar application of 1.0 kg/ ha of boron resulted in the highest fruit yield (49.01 t/ha) [61]. Boron also had a positive impact on pineapple fruit quality [62], where its application at a concentration of 2.0 mg/kg of B was deemed beneficial for enhancing fruit weight, total soluble solids (TSS), the TSS-to-acidity ratio, vitamin C content, and the concentration of aroma volatiles. As a result, the use of boron fertilizer is recommended for pineapple cultivation.

Mangoes exhibited leaf boron (B) deficiency levels ranging from 20 to 49 ppm, with sufficiency levels falling within the 50 to 100 ppm range [63]. Under the agroclimatic conditions of Doon Valley, Uttarakhand, India, an emerging physiological anomaly known as internal necrosis affects developing mango fruits due to boron deficiency, often resulting in fruit cracking. Notably, 'Dashehari' proved to be highly sensitive to both disorders, while 'Chausa' displayed the greatest tolerance. The analysis of nutrients in the leaves and fruit-bearing branches revealed that internal necrosis was primarily caused by boron deficiency. 'Dashehari' exhibited elevated leaf nitrogen levels, which may have contributed to the low levels of leaf boron and subsequently led to the occurrence of internal necrosis disorder. The most effective

solution was found to be foliar sprays of boron (in the form of disodium octaborate tetrahydrate) at a concentration of 0.10%, resulting in substantial increases in boron levels in both leaves and fruits (149.64 and 120.14% increases, respectively) of cv. 'Dashehari'. Furthermore, it was observed that the internal necrosis disorder exacerbated fruit cracking in 'Dashehari'. The study also highlighted that foliar application was more effective than soil application in terms of increasing yield while reducing internal necrosis and fruit cracking disorders [64]. In mango cv. SB Chausa, a combined application of KNO3 (1.0%) and boric acid (0.2%) led to enhanced fruit set (38–42%), increased fruit retention (56–88%), higher fruit weight, and improved yield per plant [65]. Application of Agricol at a rate of 5 grams on two sides of the plant canopy, specifically on the N-E and S-W aspects, resulted in the highest fruit yield, with each tree producing 46.2 and 45.92 kilograms of fruit [66]. Additionally, when Disodium Octaborate Tetrahydrate (DOT) was applied at a rate of 5 grams, it led to the highest number of hermaphrodite flowers per panicle (208.55 and 207.71), the best sex ratio per panicle (0.66 and 0.61), and the lowest fruit drop (51.23 and 50.50). On the other hand, the application of Agricol at 5 grams significantly improved fruit quality parameters and pulp weight (153.23 and 152.04). Maximum fruit dimensions (9.38 and 9.17 centimeters in length and width) were achieved with the use of Chemibor-P, while the application of DOT at 10 grams on the north-east and south-west canopy of the plant resulted in the highest levels of fruit bioactive compounds such as vitamin C (65.31 mg/100 g pulp and 65.64 mg/100 g pulp) and beta-carotene (3330.55 μg/100 g and 3315.18 μg/100 g). When boron was applied to mango cv. Amrapali through soil from various mineral sources, it had a clearly positive impact on the promotion of reproductive growth compared to control plants. The application of boron at two stages, namely pre-flowering and the pea stage of fruit development, influenced mango flowering and fruiting by increasing the number of flowers, reducing fruit drop, and ultimately resulting in higher mango yields. These findings underscore the role of boron in the reproductive physiology of plants, including processes like pollen tube elongation, pollen germination, sugar transport, and carbohydrate synthesis [67].

#### **7. Boron uptake and management**

Plant growth and development depends on several factors including nutrient uptake capacity and distribution to other growing parts of the plant [68]. In a single growing season crop plants may face either deficiency or toxicity [69]. This is due to very narrow range of B deficiency and toxicity in soils and plants [70] therefore, it is important to apply optimum B fertilizer for supply of B in deficient soils for normal growth, yield and quality of produce. In gladiolus, B at 0.3% showed marginal increase in flower duration while greater concentration proved toxic and lesser concentration was found to be deficient [71]. The frequency of B application would depend on doses and nature of the crop. The best response was obtained with basal application of B on crops. When boron deficiency arises, it can be addressed by applying foliar sprays containing 2.0–2.5 grams per liter of either boric acid or solubor [15]. To ensure the soil solution maintains optimal boron concentrations for maximum production, it is essential to employ environmentally friendly and advantageous techniques. These approaches not only boost boron absorption and its distribution to various plant components but also enhance soil fertility and crop yield. Some important approaches to enhance the B acquisition are as follows:

*Boron Nutrition in Horticultural Crops: Constraint Diagnosis and Their Management DOI: http://dx.doi.org/10.5772/intechopen.113367*

#### **7.1 Grafting**

Soils experiencing boron deficiency can be addressed by either incorporating boron directly into the soil or by applying boron-containing fertilizers through foliar methods. However, this approach can raise the overall cost of crop cultivation and potentially lead to boron toxicity issues due to the narrow margin between deficiency and toxicity. Therefore, an environmentally friendly and suitable alternative is the utilization of specific rootstocks tailored for different crops [72, 73]. These rootstocks possess the capability to efficiently absorb substantial amounts of boron from the soil and transport it to the upper parts of the plant, ensuring proper physiological functioning. Extensive research demonstrates that various rootstocks significantly improve the nutritional status of plants across different crops, primarily due to their effective water and mineral absorption abilities from the soil solution, surpassing those of self-rooted plants [74–77]. Furthermore, rootstocks enhance the resilience of scion cultivars to both boron deficiency [78] and toxicity [79]. The intricate physiological interactions between the scion and rootstock and their impact on mineral acquisition have been thoroughly explored in various plant species. For instance, in the case of citrus trees, Carrizo citrange (*Citrus sinensis* Osb. × *Poncirus trifoliata* [L.] Raf.) and red tangerine (*C. tangerina*) have been identified as highly effective rootstocks with strong genetic traits for boron uptake and transportation to the upper canopy under conditions of limited boron availability. Trifoliate orange (*P. trifoliata* [L.] Raf.) exhibits moderate efficiency in this regard, while sour orange (*C. aurantium* L.) and fragrant citrus (*C. medica*) have been deemed inefficient rootstocks for boron uptake and transport [72]. In studies by Liu et al. [80, 81], the impact of boron on Carrizo citrange (*C. sinensis* Osb. × *P. trifoliata* [L.] Raf.) and trifoliate orange (*P. trifoliata* [L.] Raf.) rootstocks grafted onto orange plants was examined. Their findings indicated a notable increase in boron absorption and newly absorbed boron concentration in the lower and upper leaves of Carrizo citrange grafted plants when compared to those grafted onto trifoliate orange rootstocks. In the case of pistachio trees (*Pistacia vera* cv. Kerman), *P. atlantica* rootstock demonstrated a high capacity for boron absorption and uptake of other nutrients from the soil solution, resulting in significantly higher concentrations (1.2–2.4 times more) in the leaves [82].

The distribution of ions in various plant parts can significantly differ based on their availability in the soil solution. El-Motaium et al. [79] noted a strong connection between the rootstock and boron uptake in pear plants, resulting in an increase in boron concentration of up to 50–80% in leaves and 100–300% in stems. However, the use of Prunus rootstock did not yield a notable increase in boron content in the root tissues. In the case of Newhall orange (*C. sinensis* Osb.), Sheng et al. [83] observed that when exposed to limited boron supply, boron content decreased in leaf (23–53%) and scion (40–65%) tissues but increased in rootstock parts (35–60%) when grafted onto Carrizo citrange (*C. sinensis* Osb. × *P. trifoliata* [L.] Raf.) as compared to trifoliate orange (*P. trifoliata* [L.] Raf.). Wang et al. [84] conducted a study on boron absorption patterns in four citrus rootstock-scion combinations and found that, under inadequate boron supply, the maximum boron concentration was observed in the buds and leaves of *C. sinensis* [L.] Osb. cv. Fengjie-72 when grafted onto Carrizo citrange and trifoliate orange plants. Notably, the boron accumulation in the Newhall scion grafted onto Carrizo citrange was higher (24%) compared to other combinations. Furthermore, a higher proportion of available boron (36%) was detected in the leaves of Carrizo citrange when compared to plants grafted onto trifoliate orange.

#### **7.2 Biostimulators**

Biostimulants are substances, distinct from fertilizers, soil conditioners, or pesticides, that have the capacity to impact various metabolic processes in plants, such as cell division, respiration, photosynthesis, and ion absorption, even when applied in small quantities [85]. These biostimulators can be employed to augment mineral uptake in plants while requiring minimal inputs. In recent times, biostimulants have played a pivotal role in altering plant physiology and optimizing plant growth [86]. They engage with the plant's signaling pathways to mitigate adverse plant responses during stressful conditions, ultimately promoting optimal plant development. Crops treated with biostimulants exhibit greater resilience to challenging environmental circumstances and demonstrate enhanced efficiency in ion absorption when faced with limited ion availability, primarily due to improved antioxidant production [87]. Humic substances (HS), a type of organic biostimulant, are renowned for their ability to enhance soil structure and root architecture by increasing the activity of root H + -ATPase. Consequently, they find widespread application in ion acquisition, with the effectiveness depending on factors such as concentration, plant species, and environmental conditions [88]. Field trials involving the application of biostimulants to the soil of *Vicia faba* cv. Giza beans demonstrated enhanced soil structure and ion uptake compared to untreated controls [89]. Conversely, the use of composted sewage sludge containing HS resulted in improved growth and yield of *Capsicum annuum* L. cv. Piquillo [90]. Additionally, these benefits were associated with increased availability of micronutrients in the substrate and enhanced microbial activity within plants. This microbial activity aids in reducing ion leaching by lowering soil pH through the production of organic acids like citrate, oxalate, and malate. HS forms complexes with micronutrients, and the plant's plasma membrane generates a proton motive force that facilitates the active and passive transport of ions through the symplastic pathway, thereby increasing the availability of trace elements to plants [91].

#### **7.3 Mycorrhizal fungi (MF)**

In rough lemon (*Citrus jambhiri* Lush), the application of boron through foliar spraying and soil amendment, coupled with Glomus fasciculatum inoculation, not only led to an increase in total boron accumulation in the leaves by 11–18% but also resulted in enhanced exudation of root sugars and amino acids when compared to plants that were not inoculated [92]. The presence of arbuscular mycorrhizal fungi (AMF) in the soil can impact boron concentration in plants. Research findings on the effect of AMF inoculation vary, with some studies reporting reduced boron acquisition in shoots of MF-inoculated plants [93], others showing no significant effect [94], and yet others indicating enhanced boron acquisition [95]. However, the precise role of mycorrhizal activity in relation to boron remains unproven and necessitates further investigation. The vascular structure found in higher plants underlines the importance of boron in lignification [96]. While passive uptake of boric acid seems to occur in plants, the mechanism behind mycorrhizal boron uptake is not yet fully understood. The critical role of sugar alcohols such as sucrose, sorbitol, and mannitol in the remobilization of boron within plant tissues is well-documented [4, 97]. According to Lewis's hypothesis [96], sucrose, due to its low affinity for boron in vascular plants, is primarily responsible for boron mobilization. In contrast, fungal carbohydrates, especially mannitol, exhibit a high affinity for forming complexes with boron, leading to limited boron mobility from the fungal symbiotic partner to the host plant. However,

certain mycelia have been observed to facilitate the mobility of the mannitol-boron complex, allowing for the continuous uptake and long-distance transport of boron in plants [6]. While it is established that the application of AMF can aid in boron acquisition and transport within plants, there are still specific aspects of this process that require further investigation.

#### **7.4 Plant-growth-promoting rhizobacteria (PGPR)**

Rhizobacteria, also referred to as plant-growth-promoting rhizobacteria (PGPR), are beneficial and actively root-colonizing microorganisms that establish a symbiotic relationship with plant roots. They play crucial roles in various agricultural aspects, including nitrogen fixation [98], improving tolerance to salinity and drought [99], producing enzymes that combat pathogenic microorganisms, solubilizing nutrients, and generating phytohormones like IAA, cytokinins, and gibberellins, which stimulate root growth [100]. This root proliferation, in turn, enhances water and nutrient uptake by plants.

In the case of lentils (*Lens culinaris* Medik), the inoculation of PGPR not only increased nitrogen (N) uptake (2.26–2.95%) and phosphorus (P) uptake (0.52– 0.82%) in the roots, stems, and grains but also improved plant growth parameters such as root and shoot length, as well as their fresh and dry weights. Additionally, the application of PGPR resulted in higher levels of phytohormones (IAA, GA3) and increased macro- and micronutrient concentrations in crops like *Raphanus sativus* and Musa spp. [101, 102]. In potatoes (*Solanum tuberosum* L.), the P content saw a 43.1% improvement with the introduction of the *Bacillus cereus* P31 strain, while the *Achromobacter xylosoxidans* strain P35 increased N and K content by up to 50.5 and 48.3%, respectively [103].

Numerous studies support the role of bacteria in absorbing excessive boron levels from soil solutions. These studies involve B-tolerant bacterial strains belonging to genera such as *Bacillus, Gracilibacillus, Lysinibacillus, Boronitolerans, Variovorax, Pseudomonas, Mycobacterium*, and *Rhodococcus*, which are capable of absorbing toxic levels of boron from the soil [104, 105]. Cheke et al. [106] reported that the availability of micronutrients (Fe, Mn, Cu, Zn, and B) was higher in the rhizosphere soil compared to non-rhizosphere soil, indicating that the tree rhizosphere has an impact on the availability of trace elements in the soil. In a study conducted in Nagaland, India, rhizosphere soils collected from healthy Khasi mandarin (*Citrus reticulata* Blanco) plants in an orchard displayed a higher bacterial population compared to the fungal population in the rhizosphere of four high-yielding plants [107]. However, additional research is necessary to identify efficient boron-capturing bacteria that can enhance boron availability for crops under conditions of limited boron supply.

#### **7.5 Nanotechnology**

Nanotechnology presents an innovative strategy applicable in agriculture for managing both biotic and abiotic stress, detecting diseases, and improving nutrient absorption [108, 109]. This cutting-edge technology is essential to address the challenges of limited nutrient and water resources while aiming to increase the production of high-quality agricultural crops. Nanotechnology enhances plant production and nutrient utilization efficiency by requiring fewer resources compared to traditional methods. Nanoparticles (NPs) have unique physicochemical properties that positively impact plant metabolism, leading to increased crop yield and nutritional

value [110]. To illustrate, the use of copper NPs in watermelon cultivation resulted in improved plant growth and development compared to the control group [111]. Baruah and Dutta [112] demonstrated that hydrogels and zeolites have the capacity to absorb environmental contaminants and enhance soil water retention. Chitosan NPs have proven effective in reducing fertilizer consumption, contributing to a reduction in environmental pollution [108]. While the application of CeO2 and ZnO NPs did not lead to an increase in macronutrient concentrations in *Cucumis sativus* fruit, they did elevate micronutrient levels [113]. The use of nano-titanium dioxide (TiO2) boosted chlorophyll synthesis and photosynthetic activity by enhancing ion uptake efficiency in spinach [114]. Consequently, adopting nanotechnology approaches is a promising strategy for enhancing boron uptake and utilization efficiency in plants, potentially reducing the need for boron fertilizer in crop cultivation.

#### **8. Conclusions**

Boron (B) is a vital trace element essential for the proper physiological functioning of higher plants. B deficiency represents a nutritional disorder that has detrimental effects on plant metabolism and growth. The ability of crops to efficiently utilize B resources can vary significantly, and it is crop-specific. Thus, from an agricultural perspective, there is a necessity to identify key cultivars among agronomic and horticultural crops, as well as different conditions that enable optimal utilization of available B resources, especially those that thrive under B-deficient conditions. The intricate relationship between rootstock and scion requires further in-depth studies to identify exceptional root systems, particularly those indigenous to specific regions that exhibit tolerance to both B deficiency and toxicity. Grafting and arbuscular mycorrhizal fungi (AMF) inoculation have been shown to enhance various aspects of plant physiology and nutrition, with several studies highlighting their crucial role in B uptake. Additionally, there is potential for molecular-level investigations into the role of B in plants, which could pave the way for novel strategies to enhance B stress tolerance in crops. Nanotechnology is an emerging agricultural technique designed to address plant nutrition-related challenges. The combination of these techniques has the potential to improve B uptake. Research has demonstrated that the simultaneous use of grafting and copper nanoparticles (NPs) can enhance the growth and development of watermelon by increasing ion uptake. In certain plant species, the combined inoculation of mycorrhizal fungi (MF) and plant-growth-promoting rhizobacteria (PGPR) has improved growth by augmenting water and macronutrient levels. Therefore, these existing techniques should be harnessed and further refined, considering crop-specific and location-specific factors, to achieve better outcomes and enhance B uptake and utilization in plants.

*Boron Nutrition in Horticultural Crops: Constraint Diagnosis and Their Management DOI: http://dx.doi.org/10.5772/intechopen.113367*

### **Author details**

Pauline Alila Department of Horticulture, School of Agricultural Sciences, Nagaland University, Medziphema Campus, Nagaland, India

\*Address all correspondence to: paulinealila@gmail.com

© 2023 The Author(s). Licensee IntechOpen. This chapter is distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/3.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.

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## Section 2
