**3. Main results and discussions**

To support discussions related to the applications of nanoparticles and magnetic and ferro‐ electric nanocomposites in cultures of Leishmaniasis, morphological and structural character‐ ization of nanometric materials were performed. Thus, information is obtained mainly about the interaction of the nanoparticles and nanocomposites with the biological material, cooper‐ ating in the understanding of the results.

#### **3.1. Structural and morphological nanoparticle essays**

Figure 6 presents the diffraction pattern at room temperature for KSN and NZF nanoparticles, calcined at 450 °C for 2 hours. Lines and vertical bars represent experimental data and diffrac‐ tion patterns respectively, categorized in JCPDS database: 34-0108 (KSN) and 08-0234 (NZF).

**Figure 6.** X-ray diffraction: (a) KSr2Nb5O15 phase, calcined at 450 °C, together with experimental data, columns of the identity card JCPDS-34-0108 and (b) Ni0, 5Zn0, 5Fe2O4 phase, calcined at the temperature of 450 °C, together with ex‐ perimental data, columns of the identity card JCPDS-08-0234.

As can be seen in Figure 6, and according to studies conducted previously by the authors [31], the diffraction pattern obtained for the KSN shows the typical profile of a material with shortdistance ordering (amorphous), identifying only two large sets of overlapping diffraction lines indicating that the thermal energy supplied during the heat treatment was not sufficient for obtaining a crystalline material. Relative crystallinity obtained for KSN was equal to approx‐ imately 10%, compared with the same material calcined at 1150 o C.

A diffractogram obtained for NZF phase displays a set of well-resolved diffraction lines, indicating that heat treatment was suitable for the production of a material with a high crystallinity degree, relative crystallinity of 74% when compared with the same material calcined at 650 o C. For KSN phase, the formation of a tetragonal tungsten bronze structure (TTB) with P4bm spatial group (No. 100) was identified, while for phase NZF the formation of a inverse spinel structure with space group Fd3m (No. 227) was identified. Network parameters "a", "b" and "c" obtained from KSN phase and "a" for NZF phase are equal to "a" = 12.4585 Å, "b" = "c" = 3.9423 Å and "a" = 8.394 Å, respectively. The unit cell volume is equal to V = 611.90 Å3 and V = 591.435 Å3 to KSN and NZF.

**3.1. Structural and morphological nanoparticle essays**

424 Leishmaniasis - Trends in Epidemiology, Diagnosis and Treatment

Figure 6 presents the diffraction pattern at room temperature for KSN and NZF nanoparticles, calcined at 450 °C for 2 hours. Lines and vertical bars represent experimental data and diffrac‐ tion patterns respectively, categorized in JCPDS database: 34-0108 (KSN) and 08-0234 (NZF).

**Figure 6.** X-ray diffraction: (a) KSr2Nb5O15 phase, calcined at 450 °C, together with experimental data, columns of the identity card JCPDS-34-0108 and (b) Ni0, 5Zn0, 5Fe2O4 phase, calcined at the temperature of 450 °C, together with ex‐

As can be seen in Figure 6, and according to studies conducted previously by the authors [31], the diffraction pattern obtained for the KSN shows the typical profile of a material with shortdistance ordering (amorphous), identifying only two large sets of overlapping diffraction lines indicating that the thermal energy supplied during the heat treatment was not sufficient for obtaining a crystalline material. Relative crystallinity obtained for KSN was equal to approx‐

C.

perimental data, columns of the identity card JCPDS-08-0234.

imately 10%, compared with the same material calcined at 1150 o

Average crystallite size, obtained by Scherrer's equation, was equal to 2 nm for KSN and 14.7 nm for NZF. Network microstrain (γ), calculated by Williamson-Hall equation, was equal to 0.32 for KSN and 0.05 for NZF. Structural parameters obtained in this study are in agreement with values reported in previous publications [32, 33].

Figure 7 presents transmission electron microscopy (TEM) images at a temperature of 25 °C of KSN ferroelectric and NZF paramagnetic nanoparticles, both calcined at 450 °C, where (a) and (c) are 10, 000 times magnifications while (b) and (d) are 600, 000 times magnifications. The images (b) and (d) were generated from amplifications of specific regions of the images (a) and (c).

As can be seen in Figure 7 (b) and (d), for both types of primary particles, their geometry are approximately spherical due to nucleation-type particle growth mechanism, predominant in ceramic materials and also to the principle of surface energy minimizing. Average particle diameter of strontium potassium niobate is approximately 15 nm, while the average size for a primary particle of nickel-zinc ferrite is approximately 10 nm; both values are consistent with particle diameters in the scientific literature [32, 33] and agree with average-size crystallite values. As expected, KSN particle diameter for is larger than NZF particle diameter, due to the fact that tetragonal tungsten bronze structure (23 atoms/ minimal formula, pentagonal, tetragonal and trigonal sites) has greater complexity than cubic inverse spinel-type structure (7 atoms/minimal formula, octahedral and tetrahedral sites), thus the minimum cluster size to stabilize the particle tends to be higher for KSN than for NZF. Due to the difference of stage complexity, it is also expected that as both received the same heat treatment and then the same amount of thermal energy, NZF phase would be more crystalline than KSN phase, since NZF phase requires less energy to the atoms achieve their ideal atomic positions.

According to Figure 7 (a) and (c), one can identify that both ceramic phases present clusters even at nanometric scale, due to the action of secondary forces and coalescence phenomena. For KSN phase one can identify clusters with an average size of 80 nm or approximately 112 nanoparticles/cluster and for NZF phase, agglomerates with an average size of 100 nm or around 740 nanoparticles/cluster. For both estimates, clusters with spherical shape and a closepacking bundling type were considered [34]. In principle, magnetic properties displayed by NZF nanoparticles could contribute to formation of larger clusters when compared with nonmagnetic phase clusters, as reported and discussed by E. M. A. Jamal et al., for nickel magnetic particles [35]. However, cluster formation is attributed essentially to preparation method used to synthesize ceramic nanoparticles; in this case, a chemical route.

**Figure 7.** Transmission electron microscopy (TEM) at a temperature of 25 °C, of KSN ferroelectric [(a) and (b)] and NZF paramagnetic nanoparticles [(c) and (d)], calcined at 450 °C and at different magnifications.

Images acquired by Atomic Force Microscopy (AFM) at room temperature (25 °C) for KSN ferroelectric (a) and NZF paramagnetic nanoparticles (b) calcined at 450 °C are shown in Figure 8. Details on the grain boundary and the three-dimensional nanoparticle projection are given on the right.

According to Figure 8, structures on a nanometric scale were identified for both ceramic phases, in agreement with Figure 7. Images generated from amplitude data (main figure) provide qualitative information of nanostructure shape, while images generated from elevation data (three-dimensional projection) provide significant information about surface topography. Details on the grain boundary can be obtained from deflection data of the phase angle (image positioned in the third quadrant). For KSN ferroelectric nanoparticles, Figure 8 (a), a small Utilization of Composites and Nanocomposites Based on Natural Rubber and Ceramic Nanoparticles as Control... http://dx.doi.org/10.5772/57211 427

cluster is observed in detail, with size approximately equal to 100 nm composed of nanopar‐ ticles with particle size distribution between 15 and 30 nm.

Small clusters formation is a typical feature of nanoscale materials processing using chemical routes. However, it is emphasized that the nanoparticles that compose the clusters are weakly linked together through secondary interactions of electrostatic origin. For NZF paramagnetic nanoparticles, Figure 8 (b), individual nanoparticles with approximately spherical geometry are identified, as well as the union of two or more nanoparticles by coalescence process. It is feasible to notice a particle size distribution between 25 and 40 nm for NZF phase. It should be noted that the particle size distribution for KSN and NZF is consistent with previously published work [36, 37].

#### **3.2. Nanocomposite morphological study**

Images acquired by Atomic Force Microscopy (AFM) at room temperature (25 °C) for KSN ferroelectric (a) and NZF paramagnetic nanoparticles (b) calcined at 450 °C are shown in Figure 8. Details on the grain boundary and the three-dimensional nanoparticle projection are given

**Figure 7.** Transmission electron microscopy (TEM) at a temperature of 25 °C, of KSN ferroelectric [(a) and (b)] and NZF

paramagnetic nanoparticles [(c) and (d)], calcined at 450 °C and at different magnifications.

426 Leishmaniasis - Trends in Epidemiology, Diagnosis and Treatment

According to Figure 8, structures on a nanometric scale were identified for both ceramic phases, in agreement with Figure 7. Images generated from amplitude data (main figure) provide qualitative information of nanostructure shape, while images generated from elevation data (three-dimensional projection) provide significant information about surface topography. Details on the grain boundary can be obtained from deflection data of the phase angle (image positioned in the third quadrant). For KSN ferroelectric nanoparticles, Figure 8 (a), a small

on the right.

Figure 9 presents scanning electron microscopy images obtained from the sample surface, a representation of the polymer chain and the EDX spectrum for the vulcanized natural rubber NR/KSN-1phr ferroelectric and NR/NZF-1phr magnetic nanocomposites. Magnifications used were equal to 50, 000, 50, 000, and 150, 000 times.

**Figure 9.** (a) Scanning electron microscopy images of the sample surface, polymer chain representation and EDX spec‐ trum for vulcanized natural rubber, (b) ferroelectric nanocomposite NR/KSN-1phr and (c) magnetic nanocomposite NR/NZF-1phr.

In Figure 9 (a), a satisfactory surface homogeneity was observed, indicating that a vulcaniza‐ tion system in appropriate amounts and a efficient nanocomposite-preparing system was used. White spots were noticed and indicated with white arrows. Such points may be associated with the vulcanization system, in agreement with the results obtained by XRD, particularly zinc and sulfur with submicrometer dimensions (> 250 nm). According to Figure 9 (b) and (c), it is possible to identify a high dispersion of particles and small agglomerates with dimensions on the nanometer scale, between 20 nm and 80 nm, and a particle size in the submicron range.

NR/KSN-1phr ferroelectric and NR/NZF-1phr magnetic nanocomposites. Magnifications used

**Figure 9.** (a) Scanning electron microscopy images of the sample surface, polymer chain representation and EDX spec‐ trum for vulcanized natural rubber, (b) ferroelectric nanocomposite NR/KSN-1phr and (c) magnetic nanocomposite

In Figure 9 (a), a satisfactory surface homogeneity was observed, indicating that a vulcaniza‐ tion system in appropriate amounts and a efficient nanocomposite-preparing system was used. White spots were noticed and indicated with white arrows. Such points may be associated

NR/NZF-1phr.

were equal to 50, 000, 50, 000, and 150, 000 times.

428 Leishmaniasis - Trends in Epidemiology, Diagnosis and Treatment

It is suggested that the particles and small clusters are KSN and NZF nanoparticles, in accordance with the dimensional scale, dark grayish and reddish brown color in surface and inside of the nanocomposites, respectively. In EDX spectra, peaks of C, O, S and Zn were identified and are associated with the curing system and the polymer chains. EDX percentage differences observed for S and Zn among NR, NR/KSN-1phr and NR/NZF-1phr samples refer only to the position of the investigated sample and sample time exposure to X-ray.

Low percentages of K, Sr and Nb and Fe, Ni and Zn were found for samples of NR/KSN and NR/NZF and were assigned respectively to KSN and NZF nanoparticles. The values obtained are in agreement with the amount estimated by stoichiometric calculations. The difference in surface roughness observed in Figure 9 (a) and Figures 9 (b) and (c) may be associated to the mobility difference of the natural rubber polymer chain, due to the incorporation of nanopar‐ ticles, even in small mass quantities.

Images obtained by atomic force microscopy (AFM) for vulcanized natural rubber, NR/ KSN-10phr ferroelectric nanocomposite and NR/NZF-10phr magnetic nanocomposite were performed directly on the surface of the samples, and their three-dimensional projections are presented in Figure 10, while Table 3 lists the values for NR surface roughness, NR/KSN ferroelectric and NR/NZF magnetic nanocomposites, depending on the nanoparticle concen‐ tration.


**Table 3.** Surface roughness values obtained from elevation mode AFM images, for vulcanized natural rubber (NR) and nanocomposites NR/KSN and NR/NZF.

According to Figure 10 and the data in Table 3, a satisfactory superficial homogeneity is noted for vulcanized natural rubber and both functional nanocomposites samples, suggesting that appropriate parameters and vulcanization system were used. Significant differences between the natural rubber nanocomposites were observed for surface roughness. At low nanoparticle concentrations, smaller than 3 phr, there is considerable roughness growth, followed by a reduction and stabilization of this parameter with increasing concentration of nanoparticles. This suggests that for low concentrations, local phenomena of elastomeric chain orientation as stress-induced crystallization [38, 39] can be significant.

Probably, differences in roughness identified between ferroelectric and magnetic nanocom‐ posites are due to: (i) difference in interface between the nanoparticles that generate changes in polymer chains folding, (ii) different coefficients of thermal diffusion due to different ceramic phases and (iii) different anisotropies for polymer chains mobility [39].

**Figure 10.** Images obtained using atomic force microscopy (AFM) to: (a) vulcanized natural rubber, (b) NR/KSN-10phr ferroelectric nanocomposite and (c) NR/NZF-10phr magnetic nanocomposite, performed directly on the surface of samples and their three-dimensional projections.

#### **3.3. Polymer/ceramic composites and nanocomposites as an agent of control in Leishmaniasis colonies**

Neglected diseases are illnesses that prevail not only in poverty conditions, but also contribute to the framework maintenance of economic and social inequality in the country (e.g. leishma‐ niasis, dengue, Chagas disease, schistosomiasis, leprosy and others [40]). As a result of this framework, multidisciplinary research involving materials science and biotechnology areas has gained significant strength, in order to develop new materials and methods to combat these diseases. For stimulating angiogenic processes [41] and due to its significant ability to disperse particulate fillers, natural rubber and its nanocomposites emerge as potential candidates for a new generation of bioactive agents with biocide character in biotechnology.

#### *3.3.1. Biological study: toxicity evaluation*

Probably, differences in roughness identified between ferroelectric and magnetic nanocom‐ posites are due to: (i) difference in interface between the nanoparticles that generate changes in polymer chains folding, (ii) different coefficients of thermal diffusion due to different

**Figure 10.** Images obtained using atomic force microscopy (AFM) to: (a) vulcanized natural rubber, (b) NR/KSN-10phr ferroelectric nanocomposite and (c) NR/NZF-10phr magnetic nanocomposite, performed directly on the surface of

samples and their three-dimensional projections.

ceramic phases and (iii) different anisotropies for polymer chains mobility [39].

430 Leishmaniasis - Trends in Epidemiology, Diagnosis and Treatment

Due to great demand for innovation in biotechnology, nanoparticles and nanocomposites emerge as potential candidates for a new generation of biocides, and tests that assess the toxicity of these materials compared to mammalian cells comprises an important phase of the development process biotechnology.

Figure 11 presents the results of toxicity or viability evaluation of Vero cells after 48 h incu‐ bation in the presence of ceramic nanoparticles KSr2Nb5O15 and Ni0, 5Zn0, 5Fe2O4 and their respective constituent elements, depending on particle concentration in the cellular environ‐ ment [42].

According to Figure 11 for both ceramic phases and its constituent elements, except for potassium carbonate (K2CO3), there is no statistically significant decrease in cell viability at the end of the incubation period until the maximal concentration tested in this case (1000 µg/mL), compared to cells incubated only in the culture environment.

For cells culture in contact with potassium carbonate particles, there is clearly a statistically significant reduction (P < 0.01) in cell viability for concentration equal to or greater than 62.5 µg/mL. Potassium carbonate in aqueous environment tends to dissociate, originating potas‐ sium ions (K+ ) that transform the extracellular environment, which should be hypotonic, in a highly hypertonic environment. Thus, cells pass for a excessive water-loss process through the cytoplasmic membrane and unbalances in key mechanisms for cell life maintenance, such as the sodium-potassium pump, mechanisms of nerve impulse conduction, protein synthesis and cell respiration. The combination of these processes is probably the responsible for the cell death observed for mammalian cells exposed to potassium carbonate particles. However, in KSr2Nb5O15 ferroelectric phase, potassium ions (K+ ) are isolated in the interstices of the crystallographic pentagonal structure (see Figure 4), which prevents the presence of these ions in the extracellular environment.

The results of toxicity or viability evaluation of Vero cells after 48 h incubation in the presence of vulcanized natural rubber NR/KSN ferroelectric and NR/NZF magnetic nanocomposites as a function of sample concentration in the cellular environment are shown in Figure 12. In detail, images generated by optical microscopy of cells exposed to NR/KSN-50 phr and NR/NZF-50 phr nanocomposites, and also the control sample.

**Figure 11.** Cell viability in the presence of (a) ferroelectric nanoparticles, (b) magnetic nanoparticles and their respec‐ tive constituents on the basis of the concentration of particles present in the culture environment. Vero-type mamma‐ lian cells cultured in particles presence were used.

As can be seen in Figure 12, for vulcanized natural rubber and both classes of nanocomposites, regardless of nanoparticles concentration, it is not possible to observe a statistically significant reduction in cell viability at the end of the incubation period until the maximal concentration tested (in this case 4000 µg/mL), compared to cells incubated only in the culture environment. In both images generated by optical microscopy, cells attached to the substrate are observed, indicating that cells remain biologically viable and comparing the cells exposed image to the two nanocomposites types with cells grown freely, it is not possible to identify significant morphological alterations, confirming that mammalian cells were not significantly affected due to nanocomposites presence. So as significant reductions were not identified in cell viability when mammalian cells were exposed to KSr2Nb5O15 and Ni0, 5Zn0, 5Fe2O4 nanoparti‐ cles, vulcanized natural rubber and nanocomposites, one can consider that such systems have potential for using in biological systems composed by mammalian cells.

Utilization of Composites and Nanocomposites Based on Natural Rubber and Ceramic Nanoparticles as Control... http://dx.doi.org/10.5772/57211 433

**Figure 12.** Cell viability in the presence of vulcanized natural rubber (a) NR/KSN ferroelectric and (b) NR/NZF magnetic nanocomposites as a function of sample concentration in the cell environment. In detail, images generated by optical microscopy of cells exposed and not exposed to nanocomposites. Vero-type mammalian cells were used, cultured in the presence of nanocomposites.

As can be seen in Figure 12, for vulcanized natural rubber and both classes of nanocomposites, regardless of nanoparticles concentration, it is not possible to observe a statistically significant reduction in cell viability at the end of the incubation period until the maximal concentration tested (in this case 4000 µg/mL), compared to cells incubated only in the culture environment. In both images generated by optical microscopy, cells attached to the substrate are observed, indicating that cells remain biologically viable and comparing the cells exposed image to the two nanocomposites types with cells grown freely, it is not possible to identify significant morphological alterations, confirming that mammalian cells were not significantly affected due to nanocomposites presence. So as significant reductions were not identified in cell viability when mammalian cells were exposed to KSr2Nb5O15 and Ni0, 5Zn0, 5Fe2O4 nanoparti‐ cles, vulcanized natural rubber and nanocomposites, one can consider that such systems have

**Figure 11.** Cell viability in the presence of (a) ferroelectric nanoparticles, (b) magnetic nanoparticles and their respec‐ tive constituents on the basis of the concentration of particles present in the culture environment. Vero-type mamma‐

potential for using in biological systems composed by mammalian cells.

lian cells cultured in particles presence were used.

432 Leishmaniasis - Trends in Epidemiology, Diagnosis and Treatment

**Figure 13.** Population growth kinetics of Leishmania braziliensis (LB) parasite colony exposed to samples of vulcanized natural rubber, (a) NR/KSN and (b) NR/NZF nanocomposites. In detail, morphological comparison of parasites via opti‐ cal microscopy.

Figure 13 presents the kinetics of colony population development of Leishmaniasis parasites exposed to vulcanized natural rubber samples, (a) NR/KSN ferroelectric and (b) NR/NZF magnetic nanocomposites in different nanoparticles concentrations. In detail, morphological comparison of the parasites after samples exposure.

Values for parameters maximum population density, phase duration, increase and decrease rates and also colony population of Leishmania braziliensis (LB) promastigotes exposed to samples of vulcanized natural rubber and nanocomposites are listed in Table 4.


\* Generation or double time: required time for doubling the cell population.

**Table 4.** Values for population development parameters of *Leishmania braziliensis* (LB) promastigotes colonies exposed to samples of natural rubber and vulcanized nanocomposites with different nanoparticle concentrations.

According to Figure 13 and data listed in Table 4, the increase curve evolution of LB promas‐ tigote population is similar for all samples studied, indicating that the presence of the samples did not change the colony global behavior. As expected, this evolution follows the standards of a colony of microorganisms grown in an artificial environment, being composed of three well-defined stages:

**• First stage:** denominated logarithmic phase, in which the pathogen has a large nutrient amount, conditions for their physiological maturation and mitotic cell division, a linear increase of promastigotes as a function of time is identified, and the average growth rate in this stage is higher for colonies exposed to samples. This suggests that the samples presence in the culture environment promotes the cell nutrition process;

Figure 13 presents the kinetics of colony population development of Leishmaniasis parasites exposed to vulcanized natural rubber samples, (a) NR/KSN ferroelectric and (b) NR/NZF magnetic nanocomposites in different nanoparticles concentrations. In detail, morphological

Values for parameters maximum population density, phase duration, increase and decrease rates and also colony population of Leishmania braziliensis (LB) promastigotes exposed to

> **NR/KSN 5 phr**

**Sample**

**Δ%**

14.4 9.3 -35% 9.7 -33% 13.1 -9% 12.0 -17%

15.1 17.2 +14% 16.4 +9% 13.5 -11% 11.5 -24%

30 24 -20% 24 -20% 21 -30% 15 -50%

0.3 0.51 +50% 0.48 +41% 0.4 +33% 0.4 +33%

102 108 +6% 93 -9% 75 -27% 42 -59%

36 36 0% 51 +42% 72 +100% 111 +208%

0.2 0.3 +50% 0.3 +50% 0.1 -50% 0.05 -75%

**Table 4.** Values for population development parameters of *Leishmania braziliensis* (LB) promastigotes colonies exposed to samples of natural rubber and vulcanized nanocomposites with different nanoparticle concentrations.

According to Figure 13 and data listed in Table 4, the increase curve evolution of LB promas‐ tigote population is similar for all samples studied, indicating that the presence of the samples did not change the colony global behavior. As expected, this evolution follows the standards of a colony of microorganisms grown in an artificial environment, being composed of three

**NR/NZF 1 phr**

**Δ%**

**NR/NZF 5 phr**

**Δ%**

samples of vulcanized natural rubber and nanocomposites are listed in Table 4.

**Δ%**

comparison of the parasites after samples exposure.

434 Leishmaniasis - Trends in Epidemiology, Diagnosis and Treatment

**NR/KSN 1 phr**

\* Generation or double time: required time for doubling the cell population.

**NR control**

**Parameter**

**Generation time\* (h)**

**Maximum population density (106 parasites/mL)**

**Length of logarithmic phase (h)**

**Average growth of logarithmic phase (106 cells/h mL)**

**Continuous phase duration (h)**

> **Fall phase duration (h)**

**Average rate of fall phase decrease (106 cells/h mL)**

well-defined stages:


Comparing the results for the control colony and the colony with a sample of vulcanized natural rubber samples, there is no statistically significant alterations in the of population growth kinetics, thus keeping unchanged the stages of cell development and maturation. However, for colonies exposed to nanocomposite samples having both nanoparticles, there are significant changes in microbial growth patterns. It should be mentioned that, regardless of the nanoparticle type associated with natural rubber, when the concentration of nanopar‐ ticles increases, the differences between the growth curves accentuate.

There is a progressive increase in the population of promastigotes in the logarithmic phase of the colonies exposed to NR/KSN nanocomposites (Figure 13 (a)), indicating that or KSN nanoparticles could come loose from nanocomposite surface, or something related to the interaction between the nanoparticles and the polymeric matrix is generating a change of or electronic nature significant structural proteins in the medium such that the parasites are able to ingest larger amounts of nutrients coming then to be reproduced more frequently. This hypothesis corroborates the reduction of over 30% in the generation time of the colonies.

The largest amount of immature parasites generated in logarithmic phase justifies the reduc‐ tion in hours of the stationary phase, since the presence of large parasite quantities implies in a reduction of the amount of nutrients per parasite. It is worth mentioning that the population decrease noted in fall phase is intensified with increasing nanoparticles concentration in the nanocomposite, indicating that probably the same reason that is causing changes in culture proteins, facilitating their ingestion, is also hindering the absorption of these proteins by the parasites, accelerating nutritional starvation. Comparing the morphological characteristics of the parasites exposed to NR/KSN-5phr nanocomposites with colony control parasites [43, 44], one could clearly identify the kinetoplast and nucleus cell, but no significant morphological differences were observed, confirming the similarity of the curves in Figure 13 (a).

In the case of the colonies exposed to NR/NZF nanocomposite samples (Figure 13 (b)), one can identify a linear decrease in intensity of the logarithmic phase, depending of increasing concentration of nanoparticles, indicating that the presence of such nanoparticles difficult culture protein consumption and cell division by the parasite. However, a slight reduction (lower than 20%) can still be noticeable to the generation time of the colonies. With a smaller amount of parasites in culture and limited capacity of nutritional consumption in the envi‐ ronment, there is a smaller stationary phase and a fall phase greater than that of control colonies than the sample and exposed to natural rubber samples.

Comparing the morphological characteristics of the parasites exposed to NR/NZF-5phr nanocomposites with colony control parasites [43, 44], there is a clear morphological difference in cell design. Control parasites has elongated cell bodies, while for parasites in contact with NR/NZF-5phr, the cell body has approximately a circular shape.

Whereas both types of nanoparticles have nanometric sizes, the first factor to justify the identified differences is that the sum of factors such as differences in crystallinity, surface area, micro-deformations of the crystal lattice, cell volume and especially chemical composition that generate surface characteristics particular to each nanoparticle type is responsible for the differences noted in each colony. However, intrinsic interactions between cells and magnetic/ ferroelectric nanoparticle properties, which would help to explain the high specificity exhib‐ ited by nanoparticles against leishmaniasis parasites and not against mammalian cells can not be discarded, although less likely.
