Effect of thermal treatments on the matrix components, inherent glycemic potential, and bioaccessibility of phenolics and micronutrients in pearl millet rotis.

Pearl millet (PM) is a nutri-cereal rich in various macro and micronutrients required for a balanced diet. Its grains have a unique phenolic and micronutrient composition; however, the lower bioaccessibility of nutrients and rancidity of flour during storage are the major constraints in its consumption and wide popularity. Here, to explore the effect of different thermal processing methods, i.e., hydrothermal (HT), microwave (MW), and infrared (IR) treatments, on the digestion of starch, phenolics, and microelements (Fe and Zn), an in vitro digestion model consisting of oral, gastric and intestinal digestion was applied to PM rotis. The hydrothermally treated PM roti was promising as it showed lower inherent glycemic potential (60.4%) than the untreated sample (72.4%) and less enzymatic activities associated with rancidity in PM flour. FTIR revealed an increased ratio of 1047/1022 cm-1 in the hydrothermally treated sample, reflecting the enhancement of the structurally ordered degree and compactness of starch compared to other thermal treatments. A tighter and more compact microstructure with an agglomeration of starch in the hydrothermally treated PM flour was observed by SEM. These structural changes could provide a better understanding of the lower starch digestion rate in the hydrothermally treated flour. However, HT treatment significantly (P < 0.05) reduced the bioaccessibility of phenolics (10.6%) compared to native PM rotis and slightly reduced the Fe (2%) and Zn (3.2%) bioaccessibility present in PM rotis.

Identification of Candidate Genes Regulating Drought Tolerance in Pearl Millet.

Pearl millet is an important crop of the arid and semi-arid ecologies to sustain food and fodder production. The greater tolerance to drought stress attracts us to examine its cellular and molecular mechanisms via functional genomics approaches to augment the grain yield. Here, we studied the drought response of 48 inbreds representing four different maturity groups at the flowering stage. A set of 74 drought-responsive genes were separated into five major phylogenic groups belonging to eight functional groups, namely ABA signaling, hormone signaling, ion and osmotic homeostasis, TF-mediated regulation, molecular adaptation, signal transduction, physiological adaptation, detoxification, which were comprehensively studied. Among the conserved motifs of the drought-responsive genes, the protein kinases and MYB domain proteins were the most conserved ones. Comparative in-silico analysis of the drought genes across millet crops showed foxtail millet had most orthologs with pearl millet. Of 698 haplotypes identified across millet crops, MyC2 and Myb4 had maximum haplotypes. The protein-protein interaction network identified ABI2, P5CS, CDPK, DREB, MYB, and CYP707A3 as major hub genes. The expression assay showed the presence of common as well as unique drought-responsive genes across maturity groups. Drought tolerant genotypes in respective maturity groups were identified from the expression pattern of genes. Among several gene families, ABA signaling, TFs, and signaling proteins were the prospective contributors to drought tolerance across maturity groups. The functionally validated genes could be used as promising candidates in backcross breeding, genomic selection, and gene-editing schemes in pearl millet and other millet crops to increase the yield in drought-prone arid and semi-arid ecologies.

Transcriptome Analysis of Pennisetum glaucum (L.) R. Br. Provides Insight Into Heat Stress Responses.

Pennisetum glaucum (L.) R. Br., being widely grown in dry and hot weather, frequently encounters heat stress at various stages of growth. The crop, due to its inherent capacity, efficiently overcomes such stress during vegetative stages. However, the same is not always the case with the terminal (flowering through grain filling) stages of growth, where recovery from stress is more challenging. However, certain pearl millet genotypes such as 841-B are known to overcome heat stress even at the terminal growth stages. Therefore, we performed RNA sequencing of two contrasting genotypes of pearl millet (841-B and PPMI-69) subjected to heat stress (42°C for 6 h) at flowering stages. Over 274 million high quality reads with an average length of 150 nt were generated, which were assembled into 47,310 unigenes having an average length of 1,254 nucleotides, N50 length of 1853 nucleotides, and GC content of 53.11%. Blastx resulted in the annotation of 35,628 unigenes, and functional classification showed 15,950 unigenes designated to 51 Gene Ontology terms. A total of 13,786 unigenes were allocated to 23 Clusters of Orthologous Groups, and 4,255 unigenes were distributed to 132 functional Kyoto Encyclopedia of Genes and Genomes database pathways. A total of 12,976 simple sequence repeats and 305,759 SNPs were identified in the transcriptome data. Out of 2,301 differentially expressed genes, 10 potential candidate genes were selected based on log2 fold change and adjusted p value parameters for their differential gene expression by qRT-PCR. We were able to identify differentially expressed genes unique to either of the two genotypes, and also, some DEGs common to both the genotypes were enriched. The differential expression patterns suggested that 841-B 6 h has better ability to maintain homeostasis during heat stress as compared to PPMI-69 6 h. The sequencing data generated in this study, like the SSRs and SNPs, shall serve as an important resource for the development of genetic markers, and the differentially expressed heat responsive genes shall be used for the development of transgenic crops.

Interactome of millet-based food matrices: A review.

Millets are recently being recognized as emerging food ingredients with multifaceted applications. Whole grain flours made from millets, exhibit diverse chemical compositions, starch digestibility and physicochemical properties. A food matrix can be viewed as a section of food microstructure, commonly coinciding with a physical spatial domain that interacts or imparts specific functionalities to a particular food constituent. The complex millet-based food matrices can help individuals to attain nutritional benefits due to the intricate and unique digestive properties of these foods. This review helps to fundamentally understand the binary and ternary interactions of millet-based foods. Nutritional bioavailability and bioaccessibility are also discussed based on additive, synergistic, masking, the antagonistic or neutralizing effect of different food matrix components on each other and the surrounding medium. The molecular basis of these interactions and their effect on important functional attributes like starch retrogradation, gelling, pasting, water, and oil holding capacity is also discussed.

Thermal treatments reduce rancidity and modulate structural and digestive properties of starch in pearl millet flour.

Pearl millet is a nutrient dense and gluten free cereal, however it's flour remains underutilized due to the onset of rancidity during its storage. To the best of our knowledge, processing methods, which could significantly reduce the rancidity of the pearl millet flour during storage, are non-existent. In this study, pearl millet grains were subjected to a preliminary hydro-treatment (HT). Subsequently, the hydrated grain-wet flour have undergone individual and combined thermal treatments viz., hydrothermal (HTh) and thermal near infrared rays (thNIR). Effects of these thermal treatments on the biochemical process of hydrolytic and oxidative rancidity were analyzed in stored flour. A significant (p < 0.05) decrease in the enzyme activities of lipase (47.8%), lipoxygenase (84.8%), peroxidase (98.1%) and polyphenol oxidase (100%) in HT-HTh-thNIR treated flour compared to the individual treatments was documented. Upon storage (90 days), decline of 67.84% and 66.4% of free fatty acid and peroxide contents were observed in flour under HT-HTh-thNIR treatment without altering starch and protein digestibility properties. HT-HTh treated flour exhibited the highest (7.6%) rapidly digestible starch, decreased viscosity and increased starch digestibility (67.17%). FTIR analysis of HT-HTh treated flour divulged destabilization of short-range ordered crystalline structure and altered protein structures with decreased in vitro digestibility of protein. Overall, these results demonstrated the effectiveness of combined thermal treatment of HT-HTh-thNIR in reducing rancidity and preserving the functional properties of the stored flour.

Design of experiments assisted the development of inclusion complexes of ramipril using hydrophilic carriers for enhancement of solubility and dissolution rate

Abstract The goal of the present study was to develop inclusion complexes and polymers dispersions of ramipril prepared by physical mixing, kneading, co-evaporation, and solvent evaporation methods to enhance drug solubility and dissolution rate, and thereby to reduce drug dose and side effects using selected hydrophilic carriers such as β-CD, PVP-K25, PEG 4000, and HPMC K100M. The prepared formulations were characterized for solubility and in-vitro drug release studies. The systematic optimization of formulations was performed using I-Optimal experimental design by selecting factors such as type of carriers (X1), drug: carrier ratio (X2), and method of preparation (X3), and response variables including percent yield (Y1), solubility (Y2), Carr's index (Y3) and drug release in 30 min (Y4). Mathematical modeling was carried out using a quadratic polynomial model. The inclusion complex formulation (F27) was selected as an optimized batch by numerical desirability function and graphical optimization with the help of design space. The inclusion complex prepared by the co-evaporation method showed maximum drug solubility and released in pH 6.8 phosphate buffer compared to pure and other formulations. The inclusion complex is a feasible approach to improve the solubility, dissolution rate, bioavailability, and minimization of drugs' gastrointestinal toxicity upon oral administration of ramipril.

Differential Modulation of Heat-Inducible Genes Across Diverse Genotypes and Molecular Cloning of a sHSP From Pearl Millet [Pennisetum glaucum (L.) R. Br.].

The survival, biomass, and grain yield of most of the crops are negatively influenced by several environmental stresses. The present study was carried out by using transcript expression profiling for functionally clarifying the role of genes belonging to a small heat shock protein (sHSP) family in pearl millet under high-temperature stress. Transcript expression profiling of two high-temperature-responsive marker genes, Pgcp70 and PgHSF, along with physio-biochemical traits was considered to screen out the best contrasting genotypes among the eight different pearl millet inbred lines in the seedling stage. Transcript expression pattern suggested the existence of differential response among different genotypes upon heat stress in the form of accumulation of heat shock-responsive gene transcripts. Genotypes, such as WGI 126, TT-1, TT-6, and MS 841B, responded positively toward high-temperature stress for the transcript accumulation of both Pgcp70 and PgHSF and also indicated a better growth under heat stress. PPMI-69 showed the least responsiveness to transcript induction; moreover, it supports the membrane stability index (MSI) data for scoring thermotolerance, thereby suggesting the efficacy of transcript expression profiling as a molecular-based screening technique for the identification of thermotolerant genes and genotypes at particular crop growth stages. The contrasting genotypes, such as PPMI-69 (thermosusceptible) and WGI-126 and TT-1 (thermotolerant), are further utilized for the characterization of thermotolerance behavior of sHSP by cloning a PgHSP16.97 from the thermotolerant cv. WGI-126. In addition, the investigation was extended for the identification and characterization of 28 different HSP20 genes through a genome-wide search in the pearl millet genome and an understanding of their expression pattern using the RNA-sequencing (RNA-Seq) data set. The outcome of the present study indicated that transcript profiling can be a very useful technique for high-throughput screening of heat-tolerant genotypes in the seedling stage. Also, the identified PgHSP20s genes can provide further insights into the molecular regulation of pearl millet stress tolerance, thereby bridging them together to fight against the unpredicted nature of abiotic stress.

Performance and Stability of Pearl Millet Varieties for Grain Yield and Micronutrients in Arid and Semi-Arid Regions of India.

Pearl millet [Pennisetum glaucum (L.) R. Br.] is grown under both arid and semi-arid conditions in India, where other cereals are hard to grow. Pearl millet cultivars, hybrids, and OPVs (open pollinated varieties) are tested and released by the All India Coordinated Research Project on Pearl Millet (AICRP-PM) across three zones (A1, A, and B) that are classified based on rainfall pattern. Except in locations with extreme weather conditions, hybrids dominate pearl millet growing areas, which can be attributed to hybrid vigor and the active role of the private sector. The importance of OPVs cannot be ruled out, owing to wider adaptation, lower input cost, and timely seed availability to subsidiary farmers cultivating this crop. This study was conducted to scrutinize the presently used test locations for evaluation of pearl millet OPVs across India, identify the best OPVs across locations, and determine the variation in grain Fe and Zn contents across locations in these regions. Six varieties were evaluated across 20 locations in A1 and A (pooled as A) and B zones along with three common checks and additional three zonal adapted checks in the respective zones during the 2019 rainy season. Recorded data on yield and quality traits were analyzed using genotype main effects and genotype × environment interaction biplot method. The genotype × environment (G × E) interaction was found to be highly significant for all the grain yield and agronomic traits and for both micronutrients (iron and zinc). However, genotypic effect (G) was four (productive tillers) to 49 (grain Fe content) times that of G × E interaction effect for various traits across zones that show the flexibility of OPVs. Ananthapuramu is the ideal test site for selecting pearl millet cultivars effectively for adaptation across India, while Ananthapuramu, Perumallapalle, and Gurugram can also be used as initial testing locations. OPVs MP 599 and MP 600 are identified as ideal genotypes, because they showed higher grain and fodder yields and stability compared with other cultivars. Iron and zinc concentration showed highly significant positive correlation (across environment = 0.83; p < 0.01), indicating possibility of simultaneous effective selection for both traits. Three common checks were found to be significantly low yielders than the test entries or zonal checks in individual zones and across India, indicating the potential of genetic improvement through OPVs.

Influence of different types of explants in chickpea regeneration using thidiazuron seed-priming.

A comparative regeneration of three types of explants prepared from axillary meristems, plumular apices and hypocotyls of chickpea (Cicer arietinum) was carried out using four thidiazuron (TDZ) treatment methods. The first and third ones included the short-term 20 µM TDZ pre-treatment for all three explant types followed by non-supplementation or supplementation of TDZ (4 µM) into the shoot induction medium (SIM), while the second and fourth ones lacked TDZ pre-treatment followed by non-addition or addition of 4 µM TDZ in the SIM. Axillary meristem explants produced the best results with seed pre-treatment using 20 µM TDZ without TDZ in SIM and showed the highest rate of regeneration efficiency (71.33 ± 1.5%) after 20 days. Concurrently, plumular apex explants from TDZ-primed seeds was ranked second, exhibiting a regeneration percentage of 54.33 ± 2.3% in SIM without supplementation of TDZ, whereas explants from hypocotyls generated from seeds subjected to any of the TDZ treatments were not regenerated on any SIMs after 20 days.

Lipase - The fascinating dynamics of enzyme in seed storage and germination - A real challenge to pearl millet.

Pearl millet is considered as 'nutri-cereal' because of high nutrient density of the seeds. The grain has limited use because of low keeping quality of the flour due to the activities of rancidity causing enzymes like lipase, lox, pox and PPO. Among all the enzymes, lipase is most notorious because of its robust nature and high activity under different conditions. we have identified 2180 putative transcripts showing homology with different variants of lipase precursor through transcriptome data mining (NCBI BioProject acc. no. PRJNA625418). Lipase plays dual role of facilitating the germination of seeds and deteriorating the quality of the pearl millet flour through hydrolytic rancidity. Different physiochemical methods like heat treatment, micro oven, hydrothermal, etc. have been developed to inhibit lipase activity in pearl millet flour. There is further need to develop improved processing technologies to inhibit the hydrolytic and oxidative rancidity in the floor with enhanced shelf-life.

Double-blind trial of solid lipid Boswellia serrata particles (SLBSP) vs. standardized Boswellia serrata gum extract (BSE) for osteoarthritis of knee.

Objectives The present study was planned to investigate the efficacy of SLBSP vs. standardized BSE for symptomatic knee osteoarthritis (OA) treatment. Methods It was a prospective, randomized, double-blind, double-dummy, placebo-controlled, and single-centre clinical trial for symptomatic osteoarthritis of knee. Subjects were randomized to receive SLBSP capsule+BSE Placebo or BSE tablet+SLBSP placebo for two months. Patients were allowed to take rescue analgesics (Acelofenac 100 mg). Improvement in pain and function was assessed utilizing WOMAC, VAS. Level of CTX-II in urine and serum levels of inflammatory cytokines including IL-2, IL-4, IL-6, TNF-α, and IFN-γ was measured initially and at end of treatment. Results and conclusions Western Ontario and McMaster Universities osteoarthritis index (WOMAC) and Visual Analog Scale score improved markedly in SLBSP as well as in BSE arm (p < 0.05). Difference in VAS and WOMAC scores between the two arms was not statistically significant. Most significant effect was observed in the need for rescue analgesics. SLBSP caused marked lowering of pro-inflammatory cytokines levels whereas a several fold increase was noted in the BSE arm (p < 0.05). Both groups showed marked improvement in pain, SLBSP being superior to BSE with respect to reducing the need for rescue analgesics in addition to modulating inflammatory cytokines.

Cement Augmentation of Dynamic Hip Screw to Prevent Screw Cut Out in Osteoporotic Patients with Intertrochanteric Fractures: A Case Series.

PURPOSE: The purpose of this study is to describe a method of inserting cement in the femoral head before fixation with dynamic hip screw to prevent screw cut out due to osteoporosis and to evaluate its clinical outcome in these patients. MATERIALS AND METHODS: In this prospective study, 30 patients aged 60 years and older with intertrochanteric fracture were included. Bone mineral density was measured. After reaming of the femoral head and neck with a triple reamer and polymethyl methacrylate, bone cement was introduced into the femoral head using a customized nozzle and a barrel fitted on a cement gun. A Richard screw was inserted and the plate was fixed over the femoral shaft. Patients were mobilized and clinical outcomes were rated using the Salvati and Wilson's scoring system. RESULTS: More patients included in this study were between 66 and 70 years old than any other age group. The most common fracture according to the Orthopaedic Trauma Association classification was type 31A2.2 (46.7%). The T-score was found to be -2.506±0.22 (mean±standard deviation); all patients were within the range of -2.0 to -2.8. The duration of radiological union was 13.67±1.77 weeks. Salvati and Wilson's scoring at 12 months of follow up was 30.96±4.97. The majority of patients were able to perform their normal routine activities; none experienced implant failure or screw cut out. CONCLUSION: Bone cement augmentation may effectively prevent osteoporosis-related hardware complications like screw cut out in elderly patients experiencing intertrochanteric fractures.

Open Reduction and Internal Fixation of Isolated Posterior Cruciate Ligament Avulsion Fractures: Clinical and Functional Outcome.

Purpose: The posterior cruciate ligament (PCL) presents commonly with avulsion fractures of its tibial attachment. An avulsion fracture of the PCL, if not surgically fixed, may lead to secondary changes in the knee joint. Various fixation techniques have been explored with use of lag screws, steel wires, absorbable screws, suture anchors and straddle nails. The purpose of this study was to evaluate the clinical and functional outcome of open reduction and internal fixation of tibial avulsion injuries of the PCL using cannulated cancellous screws. Materials and Methods: We performed open reduction and internal fixation using cannulated cancellous screws in 14 patients (mean age, 33.9 years) with isolated PCL avulsion injuries. Patients with a duration of injury more than 12 weeks were excluded. The minimum follow-up period was 12 months. Results were assessed radiologically and clinically. Final functional outcome was assessed using the Lysholm knee scoring system. Results: The average follow-up period was 13.5 months. At the final follow-up, all 14 patients achieved fracture union. The average flexion was 121.7°±9.2° with full extension achieved in all patients. Mild instability (1+) was noted in 4 patients. The Lysholm functional score was excellent in 11 patients, good in 2 patients and fair in 1 patient with an average score of 97±7.6. Conclusions: Stable early fixation for PCL avulsion injuries with early controlled mobilization provided excellent to good results. However, fixation should not be discouraged for late-presenting patients because good to fair results can be achieved in such cases.

Reverse Peroneal Artery Flap for Large Heel and Sole Defects: A Reliable Coverage.

BACKGROUND: Large soft tissue defects of ankle and foot always have been challenging to reconstruct. Reverse sural flaps, free flaps have been used for this problem with variable success. Reverse peroneal artery flap is an option to use with reliability without microvascular repair. Connections of peroneal artery around talus and ankle joint are deep and reliable with anterior tibial and posterior tibial artery. Arterial inflow and venous drainage improved with including short saphenous vein and reverse sural artery in the flap. METHODS: Ten patients with large defects around heel underwent reconstruction with (RPAF) reverse peroneal artery flap (pedicled) over a period of 2 years. Final inset given after 18-21 days of primary surgery. The mean age of these patients was 45 years. RESULTS: Of the 10 flaps, all showed complete survival without even marginal necrosis. Two patients had minor donor site problems that settled with conservative management. CONCLUSION: RPAF is a very reliable flap for the coverage of large soft tissue defects of the heel and sole. Large defects can be reconstructed without microvascular surgery and without compromising major vessel of foot region. If some experience with perforator flaps and free fibula is there then RPAF is easy to execute with reliability.

Deciphering Genomic Regions for High Grain Iron and Zinc Content Using Association Mapping in Pearl Millet.

Micronutrient malnutrition, especially deficiency of two mineral elements, iron [Fe] and zinc [Zn] in the developing world needs urgent attention. Pearl millet is one of the best crops with many nutritional properties and is accessible to the poor. We report findings of the first attempt to mine favorable alleles for grain iron and zinc content through association mapping in pearl millet. An association mapping panel of 130 diverse lines was evaluated at Delhi, Jodhpur and Dharwad, representing all the three pearl millet growing agro-climatic zones of India, during 2014 and 2015. Wide range of variation was observed for grain iron (32.3-111.9 ppm) and zinc (26.6-73.7 ppm) content. Genotyping with 114 representative polymorphic SSRs revealed 0.35 mean gene diversity. STRUCTURE analysis revealed presence of three sub-populations which was further supported by Neighbor-Joining method of clustering and principal coordinate analysis (PCoA). Marker-trait associations (MTAs) were analyzed with 267 markers (250 SSRs and 17 genic markers) in both general linear model (GLM) and mixed linear model (MLM), however, MTAs resulting from MLM were considered for more robustness of the associations. After appropriate Bonferroni correction, Xpsmp 2261 (13.34% R2-value), Xipes 0180 (R2-value of 11.40%) and Xipes 0096 (R2-value of 11.38%) were consistently associated with grain iron and zinc content for all the three locations. Favorable alleles and promising lines were identified for across and specific environments. PPMI 1102 had highest number (7) of favorable alleles, followed by four each for PPMFeZMP 199 and PPMI 708 for across the environment performance for both grain Fe and Zn content, while PPMI 1104 had alleles specific to Dharwad for grain Fe and Zn content. When compared with the reference genome Tift 23D2B1-P1-P5, Xpsmp 2261 amplicon was identified in intergenic region on pseudomolecule 5, while the other marker, Xipes 0810 was observed to be overlapping with aspartic proteinase (Asp) gene on pseudomolecule 3. Thus, this study can help in breeding new lines with enhanced micronutrient content using marker-assisted selection (MAS) in pearl millet leading to improved well-being especially for women and children.

Trans-differentiation of iris pigmented epithelial cells of Euphlyctis cyanophlyctis tadpoles into lens in vitro.

Meshed pigmented iris epithelium along with neural retina of tadpoles of the frog Euphlyctis cyanophlyctis were found to undergo dedifferentiation and subsequently transdifferentiate into lens in culture medium. During lag period, depigmentation (dedifferentiation) occurred in many cells. When culture became confluent 3-4 weeks after seeding tiny lens like structures differentiated from foci of cultured pigmented iris epithelium cells. The percentage of lens formation was higher in vitamin A treated cases. The culture system appears to be a suitable for investigating the changes occurred during trans-differentiation of pigmented epithelial cells into lens.