**2.1. Synthesis of supramolecular systems of water-soluble polysaccharides and their stability in aqueous solutions**

In the course of this study, different conditions of mechanochemical synthesis and various complexing agents have been tried and compared in terms of efficacy. The complexing agents were: arabinogalactan (AG), a water-soluble larch polysaccharide derived from *Larix sibirica* Ledeb. and *Larix gmelinii* (Rupr.), fibregum (FG), a glycoprotein of acacia gum, fruit pectin (PC), hydroxyethyl starch (HES200/0,5), dextrans (D) 10. 40. 70, and ß-cyclodextrin (CD), the latter chosen as standard for being widely used in pharmaceutics. The mixtures of powdered components (polysaccharides/complexing agents and drugs) were dispersed in ball mills at greater or lower intensities in laboratory planetary- and rotary-type mills, respectively. Milder rotary milling was predominantly applied because the molecular-level mixing in a planetary mill, common to laboratory studies of mechanochemical modification of drugs [9], may partly destroy the material and pose scaling problems. The materials processed by nondestructive rotary milling [10, 11, 21], instead, interact to produce solid dispersed systems of components (composite aggregates of superdispersed particles), and the process is easily scaled onto industrial flow mills.

The obtained compositions were checked for drug contents to avoid unwanted chemical reactions. Formation of supramolecular complexes was identified from changes in solubility of drugs in the water solution of the compositions [22].

Dissolution and complexing of poorly water-soluble drugs can be illustrated by such simplified equations:

$$\mathsf{Drug}\_{\mathsf{solid}} \leftarrow \succ \mathsf{Drug}\_{\mathsf{solution}} \tag{1}$$

$$\text{Drug}\_{\text{solution}} + \text{CA}\_{\text{solution}} \leftarrow \succ(\text{Drug} \cdot \text{CA})\_{\text{solution}} \tag{2}$$

Equilibrium, according to (2), is given by

574 The Complex World of Polysaccharides

properties of starting materials:

3. composite materials: aggregates of powdered particles.

micelles) that enclose drug molecules and provide their solubility.

and synthesis of polyfluorinated aromatic compounds [19, 20].

**Figure 1.** Mechanochemical transformations in mixtures of solids organic substances.

report the results of pharmacological testing.

in various mills.

medium);

components;

This difficulty may be surmountable with solid-state chemistry approaches, specifically, with mechanochemical transformations in mixtures of solids [9-11]. Unlike the liquid-phase synthesis, mechanochemical treatment is a simpler single-stage process going without solvents or melts and respective additional procedures. The flow chart in Fig. 1 shows a simplified sequence of transformations the powder mixtures experience during dry milling

There may be three types of main products relevant to our study, depending on the

1. "molecular dispersions", or solid solutions of drugs in excess filling (dispersion

2. supramolecular complexes or products of chemical reactions between the

In fact, they all are solid dispersions that form supramolecular structures (complexes or

Generally, solid-phase processes have a number of advantages in laboratory and technological uses as they yield, in a shorter time, materials which the classical liquid-phase technology can never provide and allow avoiding problems associated with melts or solvents and side reactions. The high potentiality of mechanical activation was proven in our previous studies [12-14], e.g., on quick-dissolving pharmaceutical compositions [15-18]

In this synopsis we present techniques for synthesizing supramolecular complexes of poorly soluble drugs with water-soluble polysaccharides or with glycyrrhizic acid (a plant-derived glycoside), describe physicochemical properties of their solid forms and solutions, and

$$\mathbf{K\_{DCA}} = \left[ \left( \mathbf{Drug} \cdot \mathbf{CA} \right) \text{solution} \right] / \left[ \mathbf{Drug} \text{solution} \right] \ge \left[ \mathbf{CA} \text{solution} \right] \tag{3}$$

where Drugsolid is the drug in a crystalline solid phase, in equilibrium with the solution; Drugsolution is the drug existing in the free form in the solution; CAsolution is the free complexing agent in the solution; (Drug **.** CA)solution is the complexing agent-drug complex in the solution; KDCA is the constant of supramolecular complexing.

The value Drugsolution corresponds to the thermodynamic equilibrium solubility in the absence of complexing agents. In the case of complexing, the total concentration of the dissolved drug Cdrug equals the sum its free and bound forms.

$$\text{C\_{Drug}} = \text{[Drug\_{solution}]} + \text{[(Drug\_ CA)\_{solution}]} \tag{4}$$

Thus, the solubility increase of a drug in the solution (X) in the presence of a complexing agent is

$$\mathcal{X} = \mathbb{C}\_{\text{Drug}} / \text{[Drug}\_{\text{solution}}] = 1 + \mathbb{K}\_{\text{DCA}} \cdot \text{[CA solution]} \tag{5}$$

In our view, X is a good proxy of binding strength in the supramolecular complexes drugs may form with various water-soluble polymers.

All poorly soluble drugs we studied have shown a notable solubility increase when became incorporated into compositions with complexing agents. Table 1 below shows solubility data reported in [22-32] as far as published for the first time in this review.

The binding strength in the complexes grows in the series "dextran 70 < dextrans 40 and 10 ~ < HES < ß-cyclodextrin, fibregum < pectin < arabinogalactan". Complexing of pectin with mezapam and clozapine most probably occurs by acid-base reactions, which accounts for quite a high binding strength. However, other complexing agents lack acid-base groups and the interaction mechanism is most likely "hydrophobic", as in the case of cyclodextrin complexes. Thus, the mechanochemical treatment strengthens considerably the drug binding in compositions. The solubility of drugs increases, depending on the way of mixing, in the series "mixing without milling < high-rate milling < low-rate milling.

The obtained compositions were analyzed by X-ray powder diffraction and thermal methods. All non-processed mixtures showed X-ray and thermal features typical of crystalline drugs, which disappeared or decreased markedly after milling. Therefore, drugs in the ground mixtures partly or fully loose their crystallinity, possibly, as their solid phase becomes disordered and their molecules are dispersed into the excess solid phase of complexing agents, with formation of solid solutions or supramolecular complexes. In the latter case, the solubility changes evidence that the analyzed compositions form more strongly bound complexes when form in the solid phase than in the solution.

## **2.2. Molecular dynamics and structure of arabonigalactan complexes**

AG-drug systems were investigated by 1Н NMR spectroscopy [22] for the molecular dynamics of complexes and the mobility of arabinogalactan (AG) molecule fragments. NMR relaxometry is applicable to molecular complexes as the spin-lattice and spin-spin relaxation times (T1 and T2, respectively) are highly sensitive to interactions and diffusion mobility of molecules. As a molecule becomes bound in a complex, its diffusion mobility slows down, and the proton relaxation times decrease notably. In the case of rapid complexsolution molecular exchange, the NMR signal decays according to the mono-exponential law. Otherwise, if the exchange is slower than the relaxation time, the kinetics is biexponential:

$$A(t) = P\_1 \cdot \exp(-t/T\_{21}) + P\_2 \cdot \exp(-t/T\_{22}) \tag{6}$$


may form with various water-soluble polymers.

agent is

milling.

the solution.

biexponential:

Thus, the solubility increase of a drug in the solution (X) in the presence of a complexing

In our view, X is a good proxy of binding strength in the supramolecular complexes drugs

All poorly soluble drugs we studied have shown a notable solubility increase when became incorporated into compositions with complexing agents. Table 1 below shows solubility

The binding strength in the complexes grows in the series "dextran 70 < dextrans 40 and 10 ~ < HES < ß-cyclodextrin, fibregum < pectin < arabinogalactan". Complexing of pectin with mezapam and clozapine most probably occurs by acid-base reactions, which accounts for quite a high binding strength. However, other complexing agents lack acid-base groups and the interaction mechanism is most likely "hydrophobic", as in the case of cyclodextrin complexes. Thus, the mechanochemical treatment strengthens considerably the drug binding in compositions. The solubility of drugs increases, depending on the way of mixing, in the series "mixing without milling < high-rate milling < low-rate

The obtained compositions were analyzed by X-ray powder diffraction and thermal methods. All non-processed mixtures showed X-ray and thermal features typical of crystalline drugs, which disappeared or decreased markedly after milling. Therefore, drugs in the ground mixtures partly or fully loose their crystallinity, possibly, as their solid phase becomes disordered and their molecules are dispersed into the excess solid phase of complexing agents, with formation of solid solutions or supramolecular complexes. In the latter case, the solubility changes evidence that the analyzed compositions form more strongly bound complexes when form in the solid phase than in

AG-drug systems were investigated by 1Н NMR spectroscopy [22] for the molecular dynamics of complexes and the mobility of arabinogalactan (AG) molecule fragments. NMR relaxometry is applicable to molecular complexes as the spin-lattice and spin-spin relaxation times (T1 and T2, respectively) are highly sensitive to interactions and diffusion mobility of molecules. As a molecule becomes bound in a complex, its diffusion mobility slows down, and the proton relaxation times decrease notably. In the case of rapid complexsolution molecular exchange, the NMR signal decays according to the mono-exponential law. Otherwise, if the exchange is slower than the relaxation time, the kinetics is

<sup>1</sup> 21 2 <sup>22</sup> *A*( ) exp( / ) exp( / ) *t P tT P tT* (6)

**2.2. Molecular dynamics and structure of arabonigalactan complexes** 

[CAsolution] (5)

X = CDrug /[Drugsolution] = 1 + KDCA .

data reported in [22-32] as far as published for the first time in this review.


1 – To determine the solubility of the drug, machined mixture of complexing agent/drug, in amounts of 0.4 grams, as well as the linkage of individual substances which are equivalent to their content in the above mixture was dissolved in 5 ml of water while stirring with a magnetic stirrer at +25 ° C till reaching constant concentration. The concentration of drug in the solution was analyzed by HPLC.

2 – mixing without mechanical treatment;

3 – treatment in a planetary mill, acceleration 40 g;

4 – treatment in a rotary ball mill, acceleration 1 g;

**Table 1.** Increase in water solubility of some drugs as a result of complexing.

The fast component P1 and the slow component P2 correspond, respectively, to the shares of molecules in the complex and in the solution. Typical T2 values are 0.5-1 s for molecules in the solution and 0.03-0.09 s for those bound in the complex. Shorter T21 times mean lower mobility of drug molecules in the latter case.

Similar considerations apply to the mobility within polymers when parts of a macromolecule differ in mobility, possibly, controlled by their spin and conformations.

#### *2.2.1. T2 measurements*

578 The Complex World of Polysaccharides

Nifedipine

Dihydroquercitin

Quercitin

Ibuprofen

Albendazol

Carbenazim

Simvastatin

of drug in the solution was analyzed by HPLC. 2 – mixing without mechanical treatment; 3 – treatment in a planetary mill, acceleration 40 g; 4 – treatment in a rotary ball mill, acceleration 1 g;

Hydroxyethylstarch(1/10)4 0.04/0.222 5.5 [23] Beta-cyclodextrin (1/10)3 0.04/0.60 15.1 [23] Glycyrrhizic acid (1/10)3 0.04/0.088 2.2 [29]

Arabinogalactan (1/10)3 0.18/1.24 6.9 [26] Arabinogalactan (1/20)3 0.18/2.46 13.7 [26] Glycyrrhizic acid (1/10)3 0.18/0.92 5.1 [30]

Arabinogalactan (1/10)4 0.65/3.75 5.9 [23,27] Hydroxyethylstarch(1/10)4 0.65/1.97 3.0 [23]

Arabinogalactan (1/10)3 0.019/0.21 11.6 [28] Arabinogalactan (1/20)3 0.019/1.28 71.0 [28]

Arabinogalactan (1/10)2 0.03/0.036 1.2 [29] Arabinogalactan (1/10)4 0.03/0.85 28.4 [29] Hydroxyethylstarch (1/10)4 0.03/0.08 2.6 [29] Glycyrrhizic acid (1/10)3 0.03/0.441 14.7 [29]

Arabinogalactan (1/10)4 0.003/0.174 58.0 [32] Hydroxyethylstarch(1/10)4 0.003/0.094 31.3 [32]

Arabinogalactan (1/10)4 0.009/0.146 16.2 [32] Hydroxyethylstarch(1/10)4 0.009/0.020 2.1 [32]

Glycyrrhizic acid (1/10)3 0.0012/0.314 260 This article Arabinogalactan (1/10)3 0.0012/0.044 36,7 This article

article

article

Beta-Carotene Arabinogalactan (1/40)3 < 0.001/2.65 > 2000 [25], This

Warfarin Arabinogalactan (1/40)3 0.021/0.111 5.3 [31]

Contaxantine Arabinogalactan (1/40)3 < 0.001/2.64 > 2000 [25], This

1 – To determine the solubility of the drug, machined mixture of complexing agent/drug, in amounts of 0.4 grams, as well as the linkage of individual substances which are equivalent to their content in the above mixture was dissolved in 5 ml of water while stirring with a magnetic stirrer at +25 ° C till reaching constant concentration. The concentration

**Table 1.** Increase in water solubility of some drugs as a result of complexing.

Fibregum(1/10)4 0.65/5.72 8.8 [23,27]

*Arabinogalactan* shows biexponential relaxation patterns. The calculated parameters for arabinogalactan and AG-drug complexes are listed in Table 2.


1 - T2 measurements were performed for the aromatic protons of drug molecules, to an accuracy of ± 10%

2 – solvent D2O;

3 - solvent 70% D2O + 30% CD3OD;

**Table 2.** Spin-spin relaxation times of protons for arabinogalactan and drug molecules in solutions1

The short relaxation times may correspond to the interior protons and the long times may represent the exterior protons of the polymer compound. Mechanical activation in a planetary mill increases the molecular mobility of the interior fragments but decreases their percentage. A relatively narrow ~ 6 kHz band in the 1H NMR spectra of AG powder, which stands out against a broad line associated with dipole-dipole interaction non-averagable in solids, represents a mobile phase with its integral intensity up to ~ 15% of the number of hydrogen nuclei in the sample. The mobile phase may correspond to fragments of AG macromolecules, possibly, side chains, as one may reasonably hypothesize given that water content in AG never exceeds 2 wt.%. This very fact appears to facilitate AG-Drug molecular complexing on mechanical activation of solids.

*AG-Drug systems* most often exhibit distinct biexponential kinetics as evidence that the drug molecules are either free or bound in complexes with AG. The bound molecules are more abundant and less mobile in milled samples, while the free ones keep almost invariable NMR relaxation times. The characteristic 1H NMR bands of clozapine and mezapam move to low field on complexing, possibly because the molecules become protonated at the account of minor remnant uronic acid present in AG, the shift being likewise greater in the milled samples. However, no complexing-related shifting appears in the cases of indomethacin and diazepam. The life time of drug molecules in complexes with AG must to be ~> 100 ms, judging from the conditions of slow exchange.

The system AG-diazepam offers an illustrative example. Solutions of these mixtures not subjected to mechanical treatment show mono-exponential relaxation behavior, but with shorter times than in free diazepam, likely as a result of rapid solution-complex molecular exchange. The milled mixtures, on the contrary, have biexponential kinetics corresponding to slower exchange of molecules and stronger binding.

Thus, dynamic NMR spectroscopy of all Drug-AG solutions indicates formation of supramolecular drug-polysaccharide complexes, like the data on solubility increase. Most likely, the complexing sites are at side chain spaces in the branching macromolecules. Unlike cyclodextrins, ensembles of polysaccharide molecules (including arabainogalactan) are micro-heterogeneous in mass and structure. As a result, molecular modeling of the complex is very difficult. The binding mechanism appears to lie mainly with hydrophobic interactions [33, 34] which are typical of guest-host cyclodextrin complexes. A certain support to this hypothesis comes from stronger binding of highly lipophilic drugs which are almost insoluble in water. In this case, the branched structure of AG macromolecules [35, 36] is especially favorable for complexing. However, Coulomb interactions may contribute as well in the presence of acid-base groups in polysaccharides and drugs [37].
