α‐chymotrypsin in water–acetone and water–dimethyl sulfoxide mixtures: Effect of preferential solvation and hydration

We investigated water/organic solvent sorption and residual enzyme activity to simultaneously monitor preferential solvation/hydration of protein macromolecules in the entire range of water content at 25°C. We applied this approach to estimate protein destabilization/stabilization due to the preferential interactions of bovine pancreatic α‐chymotrypsin with water‐acetone (moderate‐strength H‐bond acceptor) and water‐DMSO (strong H‐bond acceptor) mixtures. There are three concentration regimes for the dried α‐chymotrypsin. α‐Chymotrypsin is preferentially hydrated at high water content. The residual enzyme activity values are close to 100%. At intermediate water content, the dehydrated α‐chymotrypsin has a higher affinity for acetone/DMSO than for water. Residual enzyme activity is minimal in this concentration range. The acetone/DMSO molecules are preferentially excluded from the protein surface at the lowest water content, resulting in preferential hydration. The residual catalytic activity in the water‐poor acetone is ～80%, compared with that observed after incubation in pure water. This effect is very small for the water‐poor DMSO. Two different schemes are operative for the hydrated enzyme. At high and intermediate water content, α‐chymotrypsin exhibits preferential hydration. However, at intermediate water content, in contrast to the dried enzyme, the initially hydrated α‐chymotrypsin possesses increased preferential hydration parameters. At low water content, no residual enzyme activity was observed. Preferential binding of DMSO/acetone to α‐chymotrypsin was detected. Our data clearly demonstrate that the hydrogen bond accepting ability of organic solvents and the protein hydration level constitute key factors in determining the stability of protein–water–organic solvent systems.     

Gas exchange and water‐use efficiency in plant canopies

In this review, I first address the basics of gas exchange, water‐use efficiency and carbon isotope discrimination in C₃ plant canopies. I then present a case study of water‐use efficiency in northern Australian tree species. In general, C₃ plants face a trade‐off whereby increasing stomatal conductance for a given set of conditions will result in a higher CO₂ assimilation rate, but a lower photosynthetic water‐use efficiency. A common garden experiment suggested that tree species which are able to establish and grow in drier parts of northern Australia have a capacity to use water rapidly when it is available through high stomatal conductance, but that they do so at the expense of low water‐use efficiency. This may explain why community‐level carbon isotope discrimination does not decrease as steeply with decreasing rainfall on the North Australian Tropical Transect as has been observed on some other precipitation gradients. Next, I discuss changes in water‐use efficiency that take place during leaf expansion in C₃ plant leaves. Leaf phenology has recently been recognised as a significant driver of canopy gas exchange in evergreen forest canopies, and leaf expansion involves changes in both photosynthetic capacity and water‐use efficiency. Following this, I discuss the role of woody tissue respiration in canopy gas exchange and how photosynthetic refixation of respired CO₂ can increase whole‐plant water‐use efficiency. Finally, I discuss the role of water‐use efficiency in driving terrestrial plant responses to global change, especially the rising concentration of atmospheric CO₂. In coming decades, increases in plant water‐use efficiency caused by rising CO₂ are likely to partially mitigate impacts on plants of drought stress caused by global warming.     

Towards physiologically meaningful water‐use efficiency estimates from eddy covariance data

Intrinsic water‐use efficiency (iWUE) characterizes the physiological control on the simultaneous exchange of water and carbon dioxide in terrestrial ecosystems. Knowledge of iWUE is commonly gained from leaf‐level gas exchange measurements, which are inevitably restricted in their spatial and temporal coverage. Flux measurements based on the eddy covariance (EC) technique can overcome these limitations, as they provide continuous and long‐term records of carbon and water fluxes at the ecosystem scale. However, vegetation gas exchange parameters derived from EC data are subject to scale‐dependent and method‐specific uncertainties that compromise their ecophysiological interpretation as well as their comparability among ecosystems and across spatial scales. Here, we use estimates of canopy conductance and gross primary productivity (GPP) derived from EC data to calculate a measure of iWUE (G₁, “stomatal slope”) at the ecosystem level at six sites comprising tropical, Mediterranean, temperate, and boreal forests. We assess the following six mechanisms potentially causing discrepancies between leaf and ecosystem‐level estimates of G₁: (i) non‐transpirational water fluxes; (ii) aerodynamic conductance; (iii) meteorological deviations between measurement height and canopy surface; (iv) energy balance non‐closure; (v) uncertainties in net ecosystem exchange partitioning; and (vi) physiological within‐canopy gradients. Our results demonstrate that an unclosed energy balance caused the largest uncertainties, in particular if it was associated with erroneous latent heat flux estimates. The effect of aerodynamic conductance on G₁ was sufficiently captured with a simple representation. G₁ was found to be less sensitive to meteorological deviations between canopy surface and measurement height and, given that data are appropriately filtered, to non‐transpirational water fluxes. Uncertainties in the derived GPP and physiological within‐canopy gradients and their implications for parameter estimates at leaf and ecosystem level are discussed. Our results highlight the importance of adequately considering the sources of uncertainty outlined here when EC‐derived water‐use efficiency is interpreted in an ecophysiological context.     

Whole‐field macro‐ and micro‐deformation characteristic of unbound water‐loss in dentin hard tissue

High‐resolution deformation measurements in a functionally graded hard tissue such as human dentin are essential to understand the unbound water‐loss mediated changes and their role in its mechanical integrity. Yet a whole‐field, 3‐dimensional (3D) measurement and characterization of fully hydrated dentin in both macro‐ and micro‐scales remain to be a challenge. This study was conducted in 2 stages. In stage‐1, a stereo‐digital image correlation approach was utilized to determine the water‐loss and load‐induced 3D deformations of teeth in a sagittal section over consecutively acquired frames, from a fully hydrated state to nonhydrated conditions for a period up to 2 hours. The macroscale analysis revealed concentrated residual deformations at the dentin‐enamel‐junction and the apical regions of root in the direction perpendicular to the dentinal tubules. Significant difference in the localized deformation characteristics was observed between the inner and outer aspects of the root dentin. During quasi‐static loadings, further increase in the residual deformation was observed in the dentin. In stage‐2, dentin microstructural variations induced by dynamic water‐loss were assessed with environmental scanning electron microscopy and atomic force microscopy (AFM), showing that the dynamic water‐loss induced distention of dentinal tubules with concave tubular edges, and concurrent contraction of intertubular dentin with convex profile. The findings from the current macro‐ and micro‐scale analysis provided insight on the free‐water‐loss induced regional deformations and ultrastructural changes in human dentin.      

A comparative molecular analysis of water‐filled limestone sinkholes in north‐eastern Mexico

Sistema Zacatón in north‐eastern Mexico is host to several deep, water‐filled, anoxic, karstic sinkholes (cenotes). These cenotes were explored, mapped, and geochemically and microbiologically sampled by the autonomous underwater vehicle deep phreatic thermal explorer (DEPTHX). The community structure of the filterable fraction of the water column and extensive microbial mats that coat the cenote walls was investigated by comparative analysis of small‐subunit (SSU) 16S rRNA gene sequences. Full‐length Sanger gene sequence analysis revealed novel microbial diversity that included three putative bacterial candidate phyla and three additional groups that showed high intra‐clade distance with poorly characterized bacterial candidate phyla. Limited functional gene sequence analysis in these anoxic environments identified genes associated with methanogenesis, sulfate reduction and anaerobic ammonium oxidation. A directed, barcoded amplicon, multiplex pyrosequencing approach was employed to compare ～100 000 bacterial SSU gene sequences from water column and wall microbial mat samples from five cenotes in Sistema Zacatón. A new, high‐resolution sequence distribution profile (SDP) method identified changes in specific phylogenetic types (phylotypes) in microbial mats at varied depths; Mantel tests showed a correlation of the genetic distances between mat communities in two cenotes and the geographic location of each cenote. Community structure profiles from the water column of three neighbouring cenotes showed distinct variation; statistically significant differences in the concentration of geochemical constituents suggest that the variation observed in microbial communities between neighbouring cenotes are due to geochemical variation.     

Oxyma‐based phosphates for racemization‐free peptide segment couplings

Glyceroacetonide–Oxyma [(2,2‐dimethyl‐1,3‐dioxolan‐4‐yl)methyl 2‐cyano‐2‐(hydroxyimino)acetate ( 1 )] displayed remarkable physico‐chemical properties as an additive for peptide‐forming reactions. Although racemization‐free amide‐forming reactions have been established for N‐urethane‐protected α‐amino acids with EDCI, 1 , and NaHCO₃ in water or DMF‐water media, amide‐forming reactions of N‐acyl‐protected α‐amino acids and segment couplings of oligopeptides still require further development. Diethylphosphoryl–glyceroacetonide–oxyma (DPGOx 3 ) exhibits relative stability in aprotic solvents and is an effective coupling reagent for N‐acyl‐protected α‐amino acids and oligo peptide segments. The conditions reported here is also effective in lactam‐forming reactions. Unlike most of the reported coupling reagents, simple aqueous work‐up procedures can remove the reagents and by‐products generated in the reactions. Copyright © 2016 European Peptide Society and John Wiley & Sons, Ltd.     

Canopy radiation‐ and water‐use efficiencies as affected by elevated [CO2]

This study used an environmentally controlled plant growth facility, EcoCELLs, to measure canopy gas exchanges directly and to examine the effects of elevated [CO₂] on canopy radiation‐ and water‐use efficiencies. Sunflowers (Helianthus annus var. Mammoth) were grown at ambient (399 μmol mol^?1) and elevated [CO₂] (746 μmol mol^?1) for 53 days in EcoCELLs. Whole canopy carbon‐ and water‐fluxes were measured continuously during the period of the experiment. The results indicated that elevated [CO₂] enhanced daily total canopy carbon‐ and water‐fluxes by 53% and 11%, respectively, on a ground‐area basis, resulting in a 54% increase in radiation‐use efficiency (RUE) based on intercepted photosynthetic active radiation and a 26% increase in water‐use efficiency (WUE) by the end of the experiment. Canopy carbon‐ and water‐fluxes at both CO₂ treatments varied with canopy development. They were small at 22 days after planting (DAP) and gradually increased to the maxima at 46 DAP. When canopy carbon‐ and water‐fluxes were expressed on a leaf‐area basis, no effect of CO₂ was found for canopy water‐flux while elevated [CO₂] still enhanced canopy carbon‐flux by 29%, on average. Night‐time canopy carbon‐flux was 32% higher at elevated than at ambient [CO₂]. In addition, RUE and WUE displayed strong diurnal variations, high at noon and low in the morning or afternoon for WUE but opposite for RUE. This study provided direct evidence that plant canopy may consume more, instead of less, water but utilize both water and radiation more efficiently at elevated than at ambient [CO₂], at least during the exponential growth period as illustrated in this experiment.     

Partitioning of amino acids in the aqueous biphasic system containing the water‐miscible ionic liquid 1‐butyl‐3‐methylimidazolium bromide and the water‐structuring salt potassium citrate

In biotechnology, extraction by means of aqueous biphasic systems (ABS) is known as a promising tool for the recovery and purification of bio‐molecules. Over the past decade, the increasing emphasis on cleaner and environmentally benign extraction procedures has led to enhanced interest in the ABS containing ionic liquids (ILs)—a new class of non‐volatile alternative solvents. ABS composed of the hydrophilic IL {1‐butyl‐3‐methylimidazolium bromide ([C₄mim]Br)} and potassium citrate—which is easily degraded—represents a clean media to green separation of bio‐molecules. In this regard, here, the extraction capability of this ABS was evaluated through its application to the extraction of some amino acids. To gain an insight into the driving forces of amino acid partitioning in the studied IL ‐based ABS, the distribution of five model amino acids (L ‐tryptophan, L ‐phenylalanine, L ‐tyrosine, L ‐leucine, and L ‐valine) at different aqueous medium pH values and different phase compositions was investigated. The studies indicated that hydrophobic interactions were the main driving force, although electrostatic interactions and salting‐out effects were also important for the transfer of the amino acids. Moreover, based on the statistical analysis of the driving forces of amino acid partitioning in the studied IL ‐based ABS, a model was established to describe the partition coefficient of three model amino acids, L ‐tryptophan, L ‐phenylalanine, and L ‐valine, and employed to predict the partition coefficient of two other model amino acids, L ‐tyrosine and L ‐leucine. © 2011 American Institute of Chemical Engineers Biotechnol. Prog., 2011     

Isolation,purification and physicochemical characterization of water‐soluble Bacillus thuringiensis melanin

Melanins are widely used in medicine, pharmacology, cosmetics and other fields. Although several technologies for the purification of water‐insoluble dioxyphenylalanine (DOPA) melanins have been described, a source of water‐soluble melanin is highly desirable. Here we describe an effective procedure for the isolation and purification of water‐soluble melanin using the culture medium of Bacillus thuringiensis subsp. galleriae strain K1. Water‐soluble melanin from this organism has an isoelectric point (pI = 3.0–3.2) and was purified optimally by adsorbtion using the IA‐1r resin and elution as a concentrated solution. The purified melanin obtained exhibited a similar infra‐red absorbtion spectrum to synthetic melanin and contained quinolic and phenolic structures and an amino acid content of around 20% after acid hydrolysis. The molecular weight of the purified melanin determined by SDS‐PAGE was 4 kDa and the electromagnetic spin resonance spectrum of the purified microbial melanin was a slightly asymmetric singlet without hyperfine structure with about 7 Gauss width of the line between points of the maximum incline and g = 2.006. The concentration of paramagnetic centers in melanin is 0.21 × 10¹⁸ spin/g. The results obtained provide a rapid, simple and inexpensive method for the large scale purification of water soluble melanin that may have widespread applications.     

Transpiration directly regulates the emissions of water‐soluble short‐chained OVOCs

Most plant‐based emissions of volatile organic compounds are considered mainly temperature dependent. However, certain oxygenated volatile organic compounds (OVOCs) have high water solubility; thus, also stomatal conductance could regulate their emissions from shoots. Due to their water solubility and sources in stem and roots, it has also been suggested that their emissions could be affected by transport in the xylem sap. Yet further understanding on the role of transport has been lacking until present. We used shoot‐scale long‐term dynamic flux data from Scots pines (Pinus sylvestris) to analyse the effects of transpiration and transport in xylem sap flow on emissions of 3 water‐soluble OVOCs: methanol, acetone, and acetaldehyde. We found a direct effect of transpiration on the shoot emissions of the 3 OVOCs. The emissions were best explained by a regression model that combined linear transpiration and exponential temperature effects. In addition, a structural equation model indicated that stomatal conductance affects emissions mainly indirectly, by regulating transpiration. A part of the temperature's effect is also indirect. The tight coupling of shoot emissions to transpiration clearly evidences that these OVOCs are transported in the xylem sap from their sources in roots and stem to leaves and to ambient air.     

A chemiluminescence method to detect hydroquinone with water‐soluble sulphonato‐(salen)manganese(III) complex as catalyst

A water‐soluble sulphonato‐(salen)manganese(III) complex with excellent catalytic properties was synthesized and demonstrated to greatly enhance the chemiluminescence signal of the hydrogen peroxide ? luminol reaction. Coupled with flow‐injection technique, a simple and sensitive chemiluminescence method was first developed to detect hydroquinone based on the chemiluminescence system of the hydrogen peroxide–luminol–sulphonato‐(salen)manganese(III) complex. Under optimal conditions, the assay exhibited a wide linear range from 0.1 to 10 ng mL^–1 with a detection limit of 0.05 ng mL^–1 for hydroquinone. The method was applied successfully to detect hydroquinone in tap‐water and mineral‐water, with a sampling frequency of 120 times per hour. The relative standard deviation for determination of hydroquinone was less than 5.6%, and the recoveries ranged from 96.8 to 103.0%. The ultraviolet spectra, chemiluminescence spectra, and the reaction kinetics for the peroxide–luminol–sulphonato‐(salen)manganese(III) complex system were employed to study the possible chemiluminescence mechanism. The proposed chemiluminescence analysis technique is rapid and sensitive, with low cost, and could be easily extended and applied to other compounds. Copyright © 2015 John Wiley & Sons, Ltd.     

Solvent‐Polarity‐Induced Active Layer Morphology Control in Crystalline Diketopyrrolopyrrole‐Based Low Band Gap Polymer Photovoltaics

Solvent effects on the morphology of diketopyrrolopyrrole (DPP)‐based low band gap polymer (PDPPBT):phenyl‐C71‐butyric acid methyl ester (PC₇₁BM) blends are studied systematically using a mixture of a non‐aromatic polar primary solvent with high boiling point (b.p.) secondary solvents of increasing polarities. An unfavorable solvent‐PC₇₁BM interaction, due to a polarity mismatch, leads to significantly different morphology, also affecting the growth process of polymer crystallites. Non‐aromatic polar solvent produces large PC₇₁BM aggregates that increase in size with the addition of non‐polar secondary solvents. The size scales of the aggregates decrease markedly when polar solvents are instead used as the secondary solvents. This processing method fundamentally changes the behavior of phase separation, creating a percolated fibrillar type network structure. Moreover, polar secondary solvents with lower vapor pressures reduce the interfibrillar distances that enhance the device performance even more. Power conversion efficiencies (PCE) of 0.03% to 5% are obtained, depending on the solvent system used.     

Rapid and visual detection of berberine hydrochloride based on a water‐soluble pyrene derivative

In this study, a rapid method for the detection of berberine hydrochloride (BRH) was developed based on a water‐soluble pyrenyl probe, 8‐hydroxypyrene‐1,3,6‐trisulfonic acid (HPTS). This method features low cost, good selectivity, high sensitivity and a fast response. The sensing mechanism of this probe is attributed to the formation of a complex between HPTS and BRH induced by electrostatic interaction and π–π stacking. To the best of our knowledge, this is the first fluorescent sensor for BRH based on organic materials that has low cost and a visual response. The detection limit of this method was as low as 1.24 μM and the linear response range is 2–50 μM. This method also allowed rapid detection of BRH real samples.     

Solvent selection and optimization of α‐chymotrypsin‐catalyzed synthesis of N‐Ac‐Phe‐Tyr‐NH2 using mixture design and response surface methodology

A peptide, N‐Ac‐Phe‐Tyr‐NH₂, with angiotensin I‐converting enzyme (ACE) inhibitor activity was synthesized by an α‐chymotrypsin‐catalyzed condensation reaction of N‐acetyl phenylalanine ethyl ester (N‐Ac‐Phe‐OEt) and tyrosinamide (Tyr‐NH₂). Three kinds of solvents: a Tris–HCl buffer (80 mM, pH 9.0), dimethylsulfoxide (DMSO), and acetonitrile were employed in this study. The optimum reaction solvent component was determined by simplex centroid mixture design. The synthesis efficiency was enhanced in an organic‐aqueous solvent (Tris‐HCl buffer: DMSO: acetonitrile = 2:1:1) in which 73.55% of the yield of N‐Ac‐Phe‐Tyr‐NH₂ could be achieved. Furthermore, the effect of reaction parameters on the yield was evaluated by response surface methodology (RSM) using a central composite rotatable design (CCRD). Based on a ridge max analysis, the optimum condition for this peptide synthesis included a reaction time of 7.4 min, a reaction temperature of 28.1°C, an enzyme activity of 98.9 U, and a substrate molar ratio (Phe:Tyr) of 1:2.8. The predicted and the actual (experimental) yields were 87.6 and 85.5%, respectively. The experimental design and RSM performed well in the optimization of synthesis of N‐Ac‐Phe‐Tyr‐NH₂, so it is expected to be an effective method for obtaining a good yield of enzymatic peptide. © 2012 American Institute of Chemical Engineers Biotechnol. Prog., 2012     

Synthesis,spectral properties and antimicrobial activity of a new cationic water‐soluble pH‐dependent poly(propylene imine) dendrimer modified with 1,8‐naphthalimides

A three‐step synthesis was implemented to prepare a quaternary ammonium functionalized blue fluorescent poly(propylene imine) dendrimer modified with pyridinium salt of 4‐acylamino‐1,8‐naphthalimide. The new cationic dendrimer absorbs in the ultraviolet light region and emits blue fluorescence. Its spectral characteristics in organic solvents and in an aqueous solution were studied. The influence of pH on the fluorescence intensity of the dendrimer was established with regard to its use as a pH sensor. The effect of hydroxyl ions on the absorption and fluorescence spectra in dry N,N‐dimethylformamide was also investigated. The antimicrobial activity of the dendrimer was assessed against model pathogenic microorganisms in agar, liquid medium, and after its deposition on cotton fabric.     

Structural,mutagenic and in silico studies of xyloglucan fucosylation in Arabidopsis thaliana suggest a water‐mediated mechanism

The mechanistic underpinnings of the complex process of plant polysaccharide biosynthesis are poorly understood, largely because of the resistance of glycosyltransferase (GT) enzymes to structural characterization. In Arabidopsis thaliana, a glycosyl transferase family 37 (GT37) fucosyltransferase 1 (AtFUT1) catalyzes the regiospecific transfer of terminal 1,2‐fucosyl residues to xyloglucan side chains – a key step in the biosynthesis of fucosylated sidechains of galactoxyloglucan. We unravel the mechanistic basis for fucosylation by AtFUT1 with a multipronged approach involving protein expression, X‐ray crystallography, mutagenesis experiments and molecular simulations. Mammalian cell culture expressions enable the sufficient production of the enzyme for X‐ray crystallography, which reveals the structural architecture of AtFUT1 in complex with bound donor and acceptor substrate analogs. The lack of an appropriately positioned active site residue as a catalytic base leads us to propose an atypical water‐mediated fucosylation mechanism facilitated by an H‐bonded network, which is corroborated by mutagenesis experiments as well as detailed atomistic simulations.     

Improving the performance of short‐duration basmati rice in water‐saving production systems by boron nutrition

Over exploitation of groundwater and decreasing canal water resources are threating the productivity of conventional rice production systems in Asia which is the main rice bowl. Therefore, strategies are needed to produce more rice with less water in the shortest possible duration without compromising the yield to feed the increasing world population. Panicle sterility is one of the major obstacles in wide‐scale adoption of water‐saving rice production systems. Boron (B) deficiency, in water‐saving rice production systems, has been identified as a possible reason for panicle sterility. This 2‐year field study was aimed to investigate the potential of pre‐optimised boron application through various methods in improving the productivity of short‐duration basmati rice (Shaheen Basmati) in water‐saving production systems, as delivered through seed priming (0.1 mM boron), foliar spray (200 mM boron) or soil application (1 kg boron ha^?1), while hydropriming and no boron application were taken as control. Boron nutrition, by either way, improved the growth, water relations, morphology, yield‐related traits, panicle fertility, grain yield, grain quality and grain boron contents of short‐duration basmati rice; nonetheless, boron application as seed priming was superior and cost effective with maximum marginal rate of return. In conclusion, boron nutrition through seed priming is cost effective and may help improving the productivity, quality, and boron grain contents in short‐duration basmati rice under water‐saving production systems.     

High‐molecular‐weight poly(Gly‐Val‐Gly‐Val‐Pro) synthesis through microwave irradiation

In this study, we synthesized a polypeptide from its pentapeptide unit using microwave irradiation. Effective methods for polypeptide synthesis from unit peptides have not been reported. Here, we used a key elastin peptide, H‐GlyValGlyValPro‐OH (GVGVP), as the monomer peptide. It is difficult to obtain poly(Gly‐Val‐Gly‐Val‐Pro) (poly(GVGVP)) from the pentapeptide unit of elastin, GVGVP, via polycondensation. Poly(GVGVP) prepared from genetically recombinant Escherichia coli is a well‐known temperature‐sensitive polypeptide, and this temperature sensitivity is known as the lower critical solution temperature. When microwave irradiation was performed in the presence of various additives, the pentapeptide (GVGVP) polycondensation reaction proceeded smoothly, resulting in a product with a high molecular weight in a relatively good yield. The reaction conditions, like microwave irradiation, coupling agents, and solvents, were optimized to increase the reaction efficiency. The product exhibited a molecular weight greater than Mr 7000. Further, the product could be synthesized on a gram scale. The synthesized polypeptide exhibited a temperature sensitivity that was similar to that of poly(GVGVP) prepared from genetically recombinant E. coli. Therefore, this technique offers a facile and quick approach to prepare polypeptides in large amounts. Copyright © 2016 European Peptide Society and John Wiley & Sons, Ltd.     

Functional,histological and biomechanical characterization of wheat water‐mutant leaves

A wheat (Triticum turgidum subsp. durum) mutant, generated with sodium azide from wild‐type (WT) cv. ‘Trinakria’, differs in its water affinity of dry leaves, and was designated as a water‐mutant. Compared with the WT, water‐mutant leaves have lower rates of water uptake, while stomatal and cuticular transpiration do not differ. The nuclear magnetic resonance proton signals used for image reconstruction of leaf cross sections showed differences between these genotypes for the T₁ proton spin–density and the T₂ proton spin–spin relaxation time. Structural and histochemical analyses at midrib level showed that the water‐mutant has thinner leaves, with more and smaller cells per unit area of mesophyll and sclerenchyma, and has altered staining patterns of lignin and pectin‐like substances. Stress–strain curves to examine the rheological properties of the leaves showed a biphasic trend, which reveals that the tensile strength at break load and the elastic modulus of the second phase of the water‐mutant are significantly higher than for the WT. These data support the proposal of interrelationships among local biophysical properties of the leaf, the microscopic water structure, the rheological properties and the water flux rate across the leaf. This water‐mutant can be used for analysis of the genetic basis of these differences, and for identification of gene(s) that govern these traits.