Characterization of the hierarchical architecture and micromechanical properties of walnut shell (Juglans regia L.).

In the present work a comprehensive characterization of the hierarchical architecture of the walnut shell (Juglans regia L.) was carried out using scanning electron microscopy (SEM), atomic force microscopy (AFM) and confocal laser scanning microscopy (CLSM). Furthermore, micromechanical properties (hardness, HIT and elastic modulus, EIT) of plant tissues were evaluated at cell wall level by applying the instrumented indentation technique (IIT). The complex architecture of the material was described in terms of four hierarchical levels (HL): endocarp (H1), plant tissues (H2), plant cells (H3) and cell wall (H4). Our findings revealed that the walnut shell consists of a multilayer structure (sclerenchyma tissue, ST; interface tissue, IT; porous tissue, PT; and flattened parenchyma tissue, FPT), where differences in the microstructure and composition of plant tissues generate parallel gradients along the cross-section. The indentation tests showed a functional gradient with a sandwich-like configuration, i.e., a lightweight and soft layer (PT, HIT = 0.04 GPa) is located between two dense and hard layers (ST, HIT = 0.33 GPa; FPT, HIT = 0.28 GPa); where additionally there is an interface between ST and PT (IT, HIT = 0.16 GPa). This configuration is a successful strategy designed by nature to improve the protection of the kernel by increasing the strength of the shell. Therefore, the walnut shell can be considered as a functionally graded material (FGM), which can be used as bioinspiration for the design of new functional synthetic materials. In addition, we proposed some structure-property-function relationships in the whole walnut shell and in each of the plant tissues.

Effect of calcium oxalate crystals on the micromechanical properties of sclerenchyma tissue from the pecan nutshell (Carya illinoinensis).

The objective of this study was to evaluate the effect of the presence of calcium oxalate (CaOx) crystals on the micromechanical properties of sclerenchyma tissue from the pecan nutshell (Carya illinoinensis). The microstructure of the cross-section nutshell was examined using light microscopy (LM) and atomic force microscopy (AFM). Using an instrumented indentation system, indentation tests with maximum loads of 500 mN were made on the biological material where the variables studied were the number of crystals present in the evaluated area and the size of individual crystals. Microscopic analysis revealed that the pecan nutshell consists of sclerenchyma tissue with multiple CaOx crystals randomly distributed throughout the material, exhibiting prismatic shapes and various sizes. The results of the indentation tests showed that the examined areas where there were crystals (1, 2 or up to 3) presented values of hardness and elastic modulus significantly higher (P < 0.05) compared to the sclerenchyma (without crystals). Likewise, there were no significant differences (P > 0.05) between the values of the micromechanical properties of the areas evaluated as a function of the number of crystals. On the other hand, it was observed that the size of the crystals did not show a direct correlation with the mechanical properties evaluated as expected. In conclusion, the biomineralization phenomenon is a successful strategy designed by nature to improve the rigidity of the pecan nutshell, where the CaOx crystals strengthen the structure by increasing the micromechanical properties.

Effect of pepper extracts on the viability kinetics, topography and Quantitative NanoMechanics (QNM) of Campylobacter jejuni evaluated with AFM.

Campylobacter jejuni is a pathogen bacterium that causes foodborne gastroenteritis in humans. However, phenolic compounds extracted from natural sources such as capsicum pepper plant (Capsicum annuum L. var. aviculare) could inhibit the growth of C. jejuni. Therefore, different extracts were prepared using ultrasonic extraction (USE), conventional extraction (CE) and thermosonic extraction (TSE). C. jejuni was then exposed to chili extracts to examine the antimicrobial effect and their growth/death bacterial kinetics were studied using different mathematical models. Atomic force microscopy was applied to investigate the microstructural and nanomechanical changes in the bacteria. Extracts obtained by TSE had the highest phenolic content (4.59 ± 0.03 mg/g of chili fresh weight [FW]) in comparison to USE (4.12 ± 0.05 mg/g of chili FW) and CE (4.28 ± 0.07 mg/g of chili FW). The inactivation of C. jejuni was more efficient when thermosonic extract was used. The Gompertz model was the most suitable mathematical model to describe the inactivation kinetics of C. jejuni. Roughness and nanomechanical analysis performed by atomic force microscopy (AFM) provided evidence that the chili extracts had significant effects on morphology, surface, and the reduced Young's modulus of C. jejuni. The novelty of this work was integrating growth/death bacterial kinetics of C. jejuni using different mathematical models and chili extracts, and its relationship with the morphological, topographic and nanomechanical changes estimated by AFM.

Image and fractal analysis as a tool for evaluating salinity growth response between two Salicornia europaea populations.

BACKGROUND: This study describes a promising method for understanding how halophytes adapt to extreme saline conditions and to identify populations with greater resistance. Image and colour analyses have the ability to obtain many image parameters and to discriminate between different aspects in plants, which makes them a suitable tool in combination with genetic analysis to study the plants salt tolerance. To the best of our knowledge, there are no publications about the monitoring of halophytic plants by non-destructive methods for identifying the differences between plants that belong to different maternal salinity environments. The aim is to evaluate the ability of image analysis as a non-destructive method and principal component analysis (PCA) to identify the multiple responses of two S. europaea populations, and to determine which population is most affected by different salinity treatments as a preliminary model of selection. RESULTS: Image analysis was beneficial for detecting the phenotypic variability of two S. europaea populations by morphometric and colour parameters, fractal dimension (FD), projected area (A), shoot height (H), number of branches (B), shoot diameter (S) and colour change (ΔE). S was found to strongly positively correlate with both proline content and ΔE, and negatively with chlorophyll content. These results suggest that proline and ΔE are strongly linked to plant succulence, while chlorophyll decreases with increased succulence. The negative correlation between FD and hydrogen peroxide (HP) suggests that when the plant is under salt stress, HP content increases in plants causing a reduction in plant complexity and foliage growth. The PCA results indicate that the greater the stress, the more marked the differences. At 400 mM a shorter distance between the factorial scores was observed. Genetic variability analysis provided evidence of the differences between these populations. CONCLUSIONS: Our non-destructive method is beneficial for evaluating the halophyte development under salt stress. FD, S and ΔE were relevant indicators of plant architecture. PCA provided evidence that anthropogenic saline plants were more tolerant to saline stress. Furthermore, random amplified polymorphic DNA analysis provided a quick method for determining genetic variation patterns between the two populations and provided evidence of genetic differences between them.

Evaluation of Nanomechanical Properties of Tomato Root by Atomic Force Microscopy.

Here, different tissue surfaces of tomato root were characterized employing atomic force microscopy on day 7 and day 21 of growth through Young's modulus and plasticity index. These parameters provide quantitative information regarding the mechanical behavior of the tomato root under fresh conditions in different locations of the cross-section of root [cell surface of the epidermis, parenchyma (Pa), and vascular bundles (Vb)]. The results show that the mechanical parameters depend on the indented region, tissue type, and growth time. Thereby, the stiffness increases in the cell surface of epidermal tissue with increasing growth time (from 9.19 ± 0.68 to 13.90 ± 1.68 MPa) and the cell surface of Pa tissue displays the opposite behavior (from 1.74 ± 0.49 to 0.48 ± 0.55); the stiffness of cell surfaces of Vb tissue changes from 10.60 ± 0.58 to 6.37 ± 0.53 MPa, all cases showed a statistical difference (p < 0.05). Viscoelastic behavior dominates the mechanical forces in the tomato root. The current study is a contribution to a better understanding of the cell mechanics behavior of different tomato root tissues during growth.

Study of the porosity of calcified chicken eggshell using atomic force microscopy and image processing.

In this work, the porosity of the layers of calcified chicken eggshell (vertical crystal layer VCL, palisade layer PL and mammillary layer ML) was evaluated using atomic force microscopy (AFM) and image processing (IP). AFM topographic images were obtained from different locations for each layer and along the cross-section of calcified eggshell. Roughness parameters, surface area values, pore size and shape, surface porosity, area occupied by pores and pore density were obtained from AFM and IP. It was observed that the thickest layer (PL) exhibited the highest degree of porosity (surface porosity = 2.75 ± 1.68%, pore density = 162 ± 60 pores/µm2) when compared to the other two layers. In general, the pores located in all layers ("bubble pores") had circular shape and similar sizes. Measurements revealed a porosity gradient along the cross-section which varied with position, i.e., increasing surface porosity from the VCL towards the region of the PL closer to the ML, and decreasing surface porosity from this location towards the ML innermost surface. This suggests that the calcified eggshell has a sandwich-like structure where porosity may influence gas exchange and mechanical properties. The combination of AFM with IP presented here provides a simple and precise method to study porosity in calcified chicken eggshell, and this methodology could be used to examine other types of porous biological materials.

Morphological and micromechanical characterization of calcium oxalate (CaOx) crystals embedded in the pecan nutshell (Carya illinoinensis).

The morphology and micromechanical properties of the mineral crystals embedded in the pecan nutshell (Carya illinoinensis) were characterized. Qualitative and quantitative morphological analyses carried out revealed that the crystals were comprised of calcium oxalate (CaOx) and have a wide range of sizes, with prismatic shapes, distributed heterogeneously in the sclerenchyma tissue. From indentation tests, it was found that CaOx crystals are stiffer structures compared to stone cells (sclerenchyma tissue), showing hardness and elastic modulus values of 0.53 ±â€¯0.19 GPa and 9.4 ±â€¯1.80 GPa, respectively. Additionally, the values of fracture toughness (0.08 ±â€¯0.02 MPa m0.5) and the brittleness index (9336 m-0.5) revealed that these types of structures are extremely brittle. The results obtained suggest that the main function of the CaOx crystals is to provide structural support to tissue. The presented methodology demonstrates the potential of the instrumented indentation technique (IIT) for in situ micromechanical characterization of mineral crystals located in plant tissues.

Structural, mechanical and enzymatic study of pectin and cellulose during mango ripening.

Mango is an important crop worldwide, with a postharvest loss that is huge due to its climacteric behaviour. This study evaluated the softening of Tommy Atkins mangos during the ripening process. Ripening index (RPI) shown a decrease from 9.18 ±â€¯0.14 to 4.75 ±â€¯0.47. The enzymatic activity agreed with physicochemical parameters and with the structural and mechanical changes. Three pectin fractions were isolated from the mango cell wall: water soluble (WSP), chelator soluble (CSP) and diluted alkali soluble (DASP) pectin. The Younǵs modulus (E) of the primary cell wall was evaluated, it decreased from 1.69 ±â€¯1.02 to 0.39 ±â€¯0.16 MPa, which could be attributed to the softening of the fruit. A linear fit correlation between E and RPI was found. X-ray and confocal laser scanning microscopy analysis showed the changes occurred in the mango cell wall structure during maturation. Novelties of current study can be helpful in the use of mango wastes to obtain cellulose.

Physical and structural characterisation of zein and chitosan edible films using nanotechnology tools.

The use of composite edible films made from biopolymers has attracted interest as a way to reduce pollution and recycling problems; however, the relation between barrier, mechanical and structural properties of the films have been scarcely studied. The aim of this work was to evaluate composite zein-chitosan edible films by applying common nanotechnology tools and to relate the results to zein concentration and film structural changes. Rougher, more elastic, and less hard film structures with better water vapour barrier properties were obtained using larger zein concentrations. Raman spectroscopy exhibited unexpected interactions, as indicated by the disappearance of the thiol groups of cysteine in the zein films and the appearance of O=S=O and C-O-S groups in the blended materials in conjunction with the disappearance of zein ε-amino and -NH2 functional groups in the zein film samples, thereby confirming changes in the blended film structure. Zein concentration presented linear correlations with water vapour permeability (R=-0.978) and film roughness (R=0.929). The composite films presented better barrier and mechanical properties than single ingredient films. This information shows the benefit of using protein-polysaccharide blends to prepare edible films.