Synthesis of diversely substituted quinazoline-2,4(1H,3H)-diones by cyclization of tert-butyl (2-cyanoaryl)carbamates.

The synthesis of diversely substituted quinazoline-2,4(1H,3H)-diones by cyclization of tert-butyl (2-cyanoaryl)carbamates using readily accessible Boc protected o-amino nitriles is reported. The reaction proceeds smoothly at room temperature using 1 equiv. of H2O2 under basic conditions. This reaction is compatible with a variety of aromatic/heteroaromatic substrates with different functional groups. This strategy can be utilized for the simplified synthesis of goshuyuamide II and an alkaloid isolated from Zanthoxylum arborescens in good yields. This method was also applied to the synthesis of quinazoline-2,4(1H,3H)-diones that are precursors of medicinally important compounds: alfuzosin, terazosin, prazosin, IAAP, doxazosin, FK 366 (zenarestat) and KF31327.

Antioxidant and angiotensin-I-converting enzyme (ACE-I) inhibitory activities of protein hydrolysates derived from water buffalo (Bubalus bubalis) liver.

In the current study, we attempted to use ginger as a novel and natural source of protease in comparison with other commercially available enzymes to extract and characterize antioxidant and antihypertensive hydrolysates from water buffalo liver, a protein rich offal. Hydrolysis of protein extracts from buffalo liver using proteinase-K, pronase-E and ginger protease significantly increased the %degree of hydrolysis (18.5-55%) and generated low-molecular weight peptides evident from SDS-PAGE. Enzyme treated hydrolysates exhibited higher (p < 0.05) DPPH radical scavenging activity (43.7-82.4%) and angiotensin-I-converting enzyme (ACE-I) inhibitory activity (46.9-50.1%) relative to control. Mass spectrometric analysis (MALDI-TOF MS) of selected gel-filtered fractions identified few important peptides derived from nuclear ribonucleoprotein, pyruvate kinase and phosphoglycerate kinase that possess strong antioxidant activity. Present findings indicate the efficacy of partially purified ginger as a novel source of protease in generating protein hydrolysates from water buffalo liver with significant antioxidant and antihypertensive activity in vitro. We successfully demonstrated the recovery of functional bioactive peptides from water buffalo liver which presents a potential opportunity for the meat industries to economically use this important byproduct.

Needle insertion-induced quasiperiodic cone cracks in hydrogel.

Needle insertion, a standard process for various minimally invasive surgeries, results in tissue damage which sometimes leads to catastrophic outcomes. Opaqueness and inhomogeneity of the tissues make it difficult to observe the underlying damage mechanisms. In this paper, we use transparent and homogeneous polyacrylamide hydrogel as a tissue mimic to investigate the damages caused during needle insertion. The insertion force shows multiple events, characterised by a gradual increase in the force followed by a sharp fall. Synchronised recording of the needle displacement into the gel shows that each event corresponds to propagation of stable cone crack. Though sporadic uncontrolled cracking has been discussed earlier, this is the first report of nearly periodic, stable and well-controlled 3-D cone cracks inside the hydrogel during deep penetration. We show that the stress field around the needle tip is responsible for the symmetry and periodicity of the cone cracks. These results provide a better understanding of the fracture processes in soft and brittle materials and open a promising perspective in needle designs and the control of tissue damages during surgical operations.

Molecular analysis of anthocyanin biosynthesis pathway genes and their differential expression in mango peel.

Mango fruit is cherished by masses for its taste and nutrition, contributed by color, flavor, and aroma. Among these, peel color is an important trait contributing to fruit quality and market value. We attempted to elucidate the role of key genes of the anthocyanin biosynthesis pathway related to fruit peel color from the leaf transcriptome of mango cultivar Amrapali. A total of 108 mined transcript sequences were assigned to the phenylpropanoid-flavonoid pathway from which 15 contigs representing anthocyanin biosynthesis genes were annotated. Alternate splice variants were identified by mapping against genes of Citrus clementina and Vitis vinifera (closest relatives) and protein subcellular localization was determined. Phylogenetic analysis of these pathway genes clustered them into distinct groups aligning with homologous genes of Magnifera indica, C. clementina, and V. vinifera. Expression profiling revealed higher relative fold expressions in mature fruit peel of red-colored varieties (Arunika, Ambika, and Tommy Atkins) in comparison with the green-peeled Amrapali. MiCHS, MiCHI, and MiF3H alternate splice variants revealed differential gene expression. Functionally divergent variants indicate availability of an allelic pool programmed to play critical roles in peel color. This study provides insight into the molecular genetic basis of peel color and offers scope for development of biomarkers in varietal improvement programs.

Role of non-equilibrium conformations on driven polymer translocation.

One of the major theoretical methods in understanding polymer translocation through a nanopore is the Fokker-Planck formalism based on the assumption of quasi-equilibrium of polymer conformations. The criterion for applicability of the quasi-equilibrium approximation for polymer translocation is that the average translocation time per Kuhn segment, ⟨τ⟩/NK, is longer than the relaxation time τ0 of the polymer. Toward an understanding of conditions that would satisfy this criterion, we have performed coarse-grained three dimensional Langevin dynamics and multi-particle collision dynamics simulations. We have studied the role of initial conformations of a polyelectrolyte chain (which were artificially generated with a flow field) on the kinetics of its translocation across a nanopore under the action of an externally applied transmembrane voltage V (in the absence of the initial flow field). Stretched (out-of-equilibrium) polyelectrolyte chain conformations are deliberately and systematically generated and used as initial conformations in translocation simulations. Independent simulations are performed to study the relaxation behavior of these stretched chains, and a comparison is made between the relaxation time scale and the mean translocation time (⟨τ⟩). For such artificially stretched initial states, ⟨τ⟩/NK < τ0, demonstrating the inapplicability of the quasi-equilibrium approximation. Nevertheless, we observe a scaling of ⟨τ⟩ ∼ 1/V over the entire range of chain stretching studied, in agreement with the predictions of the Fokker-Planck model. On the other hand, for realistic situations where the initial artificially imposed flow field is absent, a comparison of experimental data reported in the literature with the theory of polyelectrolyte dynamics reveals that the Zimm relaxation time (τZimm) is shorter than the mean translocation time for several polymers including single stranded DNA (ssDNA), double stranded DNA (dsDNA), and synthetic polymers. Even when these data are rescaled assuming a constant effective velocity of translocation, it is found that for flexible (ssDNA and synthetic) polymers with NK Kuhn segments, the condition ⟨τ⟩/NK < τZimm is satisfied. We predict that for flexible polymers such as ssDNA, a crossover from quasi-equilibrium to non-equilibrium behavior would occur at NK ∼ O(1000).

Anomalous packing and dynamics of a polymer chain confined in a static porous environment.

Polymers in confined porous environments are ubiquitous throughout biology, physics, materials science, and engineering. Several experiments have suggested that in some porous environments, chain dynamics can become extremely slow. While phenomenological explanations exist, the exact mechanisms for these slow dynamics have not been fully characterized. In this work, we initiate a joint simulation-theory study to investigate chain packing and dynamics in a static porous environment. The main theoretical concept is the free energy of the chain partitioning into several chambers of the porous environment. Both the theoretical results and Langevin dynamics simulations show that chain packing in each of the chambers is predominantly independent of chain length; it is determined by the maximal packing of segments in each chamber. Dynamically, short chains (compared to the chamber size) become trapped in a single chamber and dynamics become extremely slow, characteristic of an Ogston sieving-like behavior. For longer chains, on the other hand, a hierarchy of slow dynamics is observed due to entropic trapping, characterized by sub-diffusive behavior and a temporary plateau in the mean square displacement. Due to the slow nature of the dynamics, the inevitable long-time diffusive behavior of the chains is not captured by our simulations. Theoretically, the slow dynamics are understood in terms of a free energy barrier required to thread the chain from one chamber to the next. There is overall qualitative and quantitative agreement between simulations and theory. This work provides foundations for a better understanding of how chain dynamics are affected by porous environments.

Single molecule electrophoresis of star polymers through nanopores: Simulations.

We study the translocation of charged star polymers through a solid-state nanopore using coarse-grained Langevin dynamics simulations, in the context of using nanopores as high-throughput devices to characterize polymers based on their architecture. The translocation is driven by an externally applied electric field. Our key observation is that translocation kinetics is highly sensitive to the functionality (number of arms) of the star polymer. The mean translocation time is found to vary non-monotonically with polymer functionality, exhibiting a critical value for which translocation is the fastest. The origin of this effect lies in the competition between the higher driving force inside the nanopore and inter-arm electrostatic repulsion in entering the pore, as the functionality is increased. Our simulations also show that the value of the critical functionality can be tuned by varying nanopore dimensions. Moreover, for narrow nanopores, star polymers above a threshold functionality do not translocate at all. These observations suggest the use of nanopores as a high-throughput low-cost analytical tool to characterize and separate star polymers.

Adsorption and encapsulation of flexible polyelectrolytes in charged spherical vesicles.

We present a theory of adsorption of flexible polyelectrolytes on the interior and exterior surfaces of a charged vesicle in an electrolyte solution. The criteria for adsorption and the density profiles of the adsorbed polymer chain are derived in terms of various characteristics of the polymer, vesicle, and medium, such as the charge density and length of the polymer, charge density and size of the vesicle, electrolyte concentration and dielectric constant of the medium. For adsorption inside the vesicle, the competition between the loss of conformational entropy and gain in adsorption energy results in two kinds of encapsulated states, depending on the strength of the polymer-vesicle interaction. By considering also the adsorption from outside the vesicle, we derive the entropic and energy contributions to the free energy change to transfer an adsorbed chain in the interior to an adsorbed chain on the exterior. In this paper, we have used the Wentzel-Kramers-Brillouin (WKB) method to solve the equation for the probability distribution function of the chain. The present WKB results are compared with the previous results based on variational methods. The WKB and variational results are in good agreement for both the interior and exterior states of adsorption, except in the zero-salt limit for adsorption in the exterior region. The adsorption criteria and density profiles for both the interior and exterior states are presented in terms of various experimentally controllable variables. Calculation of the dependencies of free energy change to transfer an adsorbed chain from the interior to the exterior surface on salt concentration and vesicle radius shows that the free energy penalty to expel a chain from a vesicle is only of the order of thermal energy.

Communication: Theory of melt-memory in polymer crystallization.

Details of crystallization processes of a polymer at the crystallization temperature Tc from its melt kept initially at the melt temperature Tm depend profoundly on the nature of the initial melt state and often are accompanied by memory effects. This phenomenon is in contrast to small molecular systems where the supercooling (Tm (0)-Tc), with Tm (0) being the equilibrium melting temperature, and not (Tm - Tc), determines the nature of crystallization. In addressing this five-decade old puzzle of melt-memory in polymer crystallization, we present a theory to describe melt-memory effects, by invoking an intermediate inhomogeneous melt state in the pathway between the melt and crystalline states. Using newly introduced dissolution temperature T1 (0) for the inhomogeneous melt state and the transition temperature Tt (0) for the transition between the inhomogeneous melt and crystalline states, analytical formulas are derived for the nucleation rate as a function of the melt temperature. The theory is general to address different kinds of melt-memory effects depending on whether Tm is higher or lower than Tm (0). The derived results are in qualitative agreement with known experimental data, while making predictions for further experiments on melt-memory.

Ratchet rectification effect on the translocation of a flexible polyelectrolyte chain.

We report a three dimensional Langevin dynamics simulation of a uniformly charged flexible polyelectrolyte chain, translocating through an asymmetric narrow channel with periodically varying cross sections under the influence of a periodic external electric field. When reflection symmetry of the channel is broken, a rectification effect is observed with a favored direction for the chain translocation. For a given volume of the channel unit and polymer length, the rectification occurs below a threshold frequency of the external periodic driving force. We have also observed that the extent of the rectification varies non-monotonically with increasing molecular weight and the strength of geometric asymmetry of the channel. Observed non-monotonicity of the rectification performance has been interpreted in terms of a competition between two effects arising from the channel asymmetry and change in conformational entropy. An analytical model is presented with predictions consistent with the simulation results.

Stochastic resonance during a polymer translocation process.

We have studied the occurrence of stochastic resonance when a flexible polymer chain undergoes a single-file translocation through a nano-pore separating two spherical cavities, under a time-periodic external driving force. The translocation of the chain is controlled by a free energy barrier determined by chain length, pore length, pore-polymer interaction, and confinement inside the donor and receiver cavities. The external driving force is characterized by a frequency and amplitude. By combining the Fokker-Planck formalism for polymer translocation and a two-state model for stochastic resonance, we have derived analytical formulas for criteria for emergence of stochastic resonance during polymer translocation. We show that no stochastic resonance is possible if the free energy barrier for polymer translocation is purely entropic in nature. The polymer chain exhibits stochastic resonance only in the presence of an energy threshold in terms of polymer-pore interactions. Once stochastic resonance is feasible, the chain entropy controls the optimal synchronization conditions significantly.

Simulation of self-assembly of polyzwitterions into vesicles.

Using the Langevin dynamics method and a coarse-grained model, we have studied the formation of vesicles by hydrophobic polymers consisting of periodically placed zwitterion side groups in dilute salt-free aqueous solutions. The zwitterions, being permanent charge dipoles, provide long-range electrostatic correlations which are interfered by the conformational entropy of the polymer. Our simulations are geared towards gaining conceptual understanding in these correlated dipolar systems, where theoretical calculations are at present formidable. A competition between hydrophobic interactions and dipole-dipole interactions leads to a series of self-assembled structures. As the spacing d between the successive zwitterion side groups decreases, single chains undergo globule → disk → worm-like structures. We have calculated the Flory-Huggins χ parameter for these systems in terms of d and monitored the radius of gyration, hydrodynamic radius, spatial correlations among hydrophobic and dipole monomers, and dipole-dipole orientational correlation functions. During the subsequent stages of self-assembly, these structures lead to larger globules and vesicles as d is decreased up to a threshold value, below which no large scale morphology forms. The vesicles form via a polynucleation mechanism whereby disk-like structures form first, followed by their subsequent merger.

Comparative evaluation of carcass traits and meat quality in native Aseel chickens and commercial broilers.

A comprehensive study was conducted to analyse the meat quality attributes, composition and carcass traits in Aseel chickens and commercial broilers at market age on the basis of physiological age. A total of 20 Aseel (26 and 56 weeks) and 20 broiler (6 weeks) chickens were divided into two groups on a live weight basis, i.e. large (≥2.5 kg) and small (<2.5 kg) with 10 birds in each subgroup. The pH of meat did not show any significant variation between Aseel and broiler chickens. The meat from heavier birds had significantly higher pH. Shear force value and hydroxyproline contents were significantly higher in Aseel chickens. Aseel birds had significantly higher red (a*) colouration and lower lightness (L*) than broiler chickens. The texture and acceptability of Aseel meat were significantly higher. Scanning electron microscopy revealed that muscle fibres in Aseels were arranged in a more coiled pattern making the muscle tough. A larger amount of connective tissue was also observed between the muscle fibres compared with the broiler chickens. The dressing percentage was significantly higher in larger chickens. Commercial broilers recorded significantly higher meat proportion and lower proportion of bone. The meat:bone ratio was 1.07:1.0 in Aseel and 1.31-1.0 in broiler chicken. Breast muscle content was significantly lower in smaller Aseel chickens. Aseel chicken had stronger and heavier backs and shanks. Abdominal fat percentage was significantly lower in Aseel (0.73-0.78%). The study concluded that the firm texture of Aseel meat was due to the high collagen content and interlocking connective tissue between the muscle fibres. The texture and acceptability of Aseel meat was higher. Aseel cocks had strong legs, lean meat and less abdominal fat, making them a high-value meat bird in addition to their aggressive fighting ability.

Molecular and morphological diversity in locally grown non-commercial (heirloom) mango varieties of North India.

Mango (Mangifera indica L.) has been cultivated and conserved in different agro-ecologies including Malihabad region in northern part of India, that is well known for housing diverse types (heirloom and commercial varieties). In the present study, 37 mango types comprising of 27 heirloom varieties from Malihabad region and 10 commercial varieties grown in North and Eastern India were assessed for morphological attributes and molecular diversity. The employed SSR markers amplified 2-13 alleles individually, cumulatively amplifying 124 alleles. These were studied for allelic diversity and genetic dissimilarity ranged from 0.035 to 0.892 arranging the varieties in three major clusters. The results revealed that majority of unique heirloom mangoes from Malihabad were different from the eastern part of the country. It is interesting to note Dashehari, a commercial variety from Malihabad was not aligned with heirloom varieties. Commercial varieties like Gulabkhas and Langra were placed in a separate group including Bombay Green, Himsagar, Dashehari, etc., indicating their dissimilarity with heirloom varieties at molecular level and thus, indicating importance for later from conservation point of view. Furthermore, the hierarchical clustering of varieties based on fruit morphology, assembled these into four groups largely influenced by fruit size. The maximum agreement subtree indicated seemingly good fit as thirteen varieties were arrayed in common grouping pattern. Appreciable dissimilarity among the heirloom varieties demonstrated by molecular analysis, underlines the importance for their on-farm conservation.

Modeling competitive substitution in a polyelectrolyte complex.

We have simulated the invasion of a polyelectrolyte complex made of a polycation chain and a polyanion chain, by another longer polyanion chain, using the coarse-grained united atom model for the chains and the Langevin dynamics methodology. Our simulations reveal many intricate details of the substitution reaction in terms of conformational changes of the chains and competition between the invading chain and the chain being displaced for the common complementary chain. We show that the invading chain is required to be sufficiently longer than the chain being displaced for effecting the substitution. Yet, having the invading chain to be longer than a certain threshold value does not reduce the substitution time much further. While most of the simulations were carried out in salt-free conditions, we show that presence of salt facilitates the substitution reaction and reduces the substitution time. Analysis of our data shows that the dominant driving force for the substitution process involving polyelectrolytes lies in the release of counterions during the substitution.

Application of response surface methodology for optimization of polygalacturonase production by Aspergillus niger.

Polygalacturonase (PG) degrades pectin into D-galacturonic acid monomers and is used widely in food industry especially for juice clarification. In the present study,. fermentation conditions for polygalacturonase production by Asgergillus niger NAIMCCF-02958, using mango peel as substrate, were optimized using the 2(3) factorial design with central composite rotatable experimental design (CCRD) of response surface methodology (RSM). The maximum PG activity 723.66 U g(-1) was achieved under pH 4.0, temperature 30 degrees C and 2% inoculum by response surface curve. The experimental value of PG activity wkas higher 607.65 U g(-1) than the predicted value 511.75 U g(-1). Under the proposed optimized conditions, the determination coefficient (R2) was equal to 0.66 indicating that the model could explain 66% of the total variation as well as establish the relationship between the variables and the responses. ANOVA analysis and the three dimensional plots also confirmed interactions among the parameters.

Communication: Charge, diffusion, and mobility of proteins through nanopores.

Implementation of Einstein's law connecting charge, diffusion coefficient, and mobility to interpret experimental data on proteins from single molecule electrophoresis through nanopores faces serious difficulties. The protein charge and diffusion coefficient, inferred with the Einstein law, can be orders of magnitude smaller than their bare values depending on the electrolyte concentration, pore diameter, chemical nature of the pore wall, and the externally applied voltage. The main contributors to the discrepancies are the coupled dynamics of the protein and its counterion cloud, confinement effects inside the pore, and the protein-pore-surface interaction. We have addressed these ingredients by harking on classical theories of electrophoresis of macroions and hydrodynamics inside pores, and deriving new results for pore-protein interactions. Putting together various components, we present approximate analytical formulas for the effective charge, diffusion coefficient, and mobility of a protein in the context of single molecule electrophoresis experiments. For the omnipresent pore-protein interactions, nonlinear dependence of the velocity of protein on voltage sets in readily and analytical formulas for this effect are presented. The derived formulas enable the determination of the bare charge and size of a protein from the experimentally measured apparent values.

Effect of charge patterns along a solid-state nanopore on polyelectrolyte translocation.

We investigate the effectiveness of charge patterns along a nanopore on translocation dynamics of a flexible polyelectrolyte. We perform a three dimensional Langevin dynamics simulation of a uniformly charged flexible polyelectrolyte translocating under uniform external electric field through a solid-state nanopore. We maintain the total charge along the pore to be constant, while varying its distribution by placing alternate charged and uncharged sections of different lengths along the pore length. Longest average translocation time is observed for a pattern corresponding to an optimum section length, with a major delay in the translocation time during the pore ejection stage. This optimum section length is independent of lengths of polyelectrolyte and pore within the range studied. A theory based on the Fokker-Planck formalism is found to successfully describe the observed trends with reasonable quantitative agreement.

Electrophoretic mobilities of counterions and a polymer in cylindrical pores.

We have simulated the transport properties of a uniformly charged flexible polymer chain and its counterions confined inside cylindrical nanopores under an external electric field. The hydrodynamic interaction is treated by describing the solvent molecules explicitly with the multiparticle collision dynamics method. The chain consisting of charged monomers and the counterions interact electrostatically with themselves and with the external electric field. We find rich behavior of the counterions around the polymer under confinement in the presence of the external electric field. The mobility of the counterions is heterogeneous depending on their location relative to the polymer. The adsorption isotherm of the counterions on the polymer depends nonlinearly on the electric field. As a result, the effective charge of the polymer exhibits a sigmoidal dependence on the electric field. This in turn leads to a nascent nonlinearity in the chain stretching and electrophoretic mobility of the polymer in terms of their dependence on the electric field. The product of the electric field and the effective polymer charge is found to be the key variable to unify our simulation data for various polymer lengths. Chain extension and the electrophoretic mobility show sigmoidal dependence on the electric field, with crossovers from the linear response regime to the nonlinear regime and then to the saturation regime. The mobility of adsorbed counterions is nonmonotonic with the electric field. For weaker and moderate fields, the adsorbed counterions move with the polymer and at higher fields they move opposite to the polymer's direction. We find that the effective charge and the mobility of the polymer decrease with a decrease in the pore radius.

Colour, myoglobin denaturation and storage stability of raw and cooked mutton chops at different end point cooking temperature.

In our study effect of different end point temperature (51 °C, 65 °C, 71 °C and 79 °C) on physicochemical and storage stability of mutton chops were evaluated. The L* (lightness) value and b* (yellowness) increased (P < 0.05) in cooked mutton chops than the raw mutton. The a* value (redness) decreased (P < 0.05) as end point temperature increased. As internal cooking temperature increased soluble myoglobin content decreased with a corresponding increase in percent myoglobin denatured. Raw mutton chops (uncooked) had lower level of oxidation (less TBA values) than cooked mutton irrespective of storage length. Initial APC of raw and cooked mutton chops ranged from log 1.75 to log 3.73 and was lower in higher end point cooking temperature. It can be concluded that as end point temperature increased, mutton chops appear less red and raw mutton had lower level of oxidation than cooked mutton chops.