Catalytic, Theoretical, and Biological Investigations of Ternary Metal (II) Complexes Derived from L-Valine-Based Schiff Bases and Heterocyclic Bases.

A new series of ternary metal complexes, including Co(II), Ni(II), Cu(II), and Zn(II), were synthesized and characterized by elemental analysis and diverse spectroscopic methods. The complexes were synthesized from respective metal salts with Schiff's-base-containing amino acids, salicylaldehyde derivatives, and heterocyclic bases. The amino acids containing Schiff bases showed promising pharmacological properties upon complexation. Based on satisfactory elemental analyses and various spectroscopic techniques, these complexes revealed a distorted, square pyramidal geometry around metal ions. The molecular structures of the complexes were optimized by DFT calculations. Quantum calculations were performed with the density functional method for which the LACVP++ basis set was used to find the optimized molecular structure of the complexes. The metal complexes were subjected to an electrochemical investigation to determine the redox behavior and oxidation state of the metal ions. Furthermore, all complexes were utilized for catalytic assets of a multi-component Mannich reaction for the preparation of -amino carbonyl derivatives. The synthesized complexes were tested to determine their antibacterial activity against E. coli, K. pneumoniae, and S. aureus bacteria. To evaluate the cytotoxic effects of the Cu(II) complexes, lung cancer (A549), cervical cancer (HeLa), and breast cancer (MCF-7) cells compared to normal cells, cell lines such as human dermal fibroblasts (HDF) were used. Further, the docking study parameters were supported, for which it was observed that the metal complexes could be effective in anticancer applications.

Mobile group II intron based gene targeting in Lactobacillus plantarum WCFS1.

The usage of recombinant lactic acid bacteria for delivery of therapeutic proteins to the mucosa has been emerging. In the present study, an attempt was made to engineer a thyA mutant of Lactobacillus plantarum (L. plantarum) using lactococcal group II intron Ll.LtrB for the development of biologically contained recombinant L. plantarum for prevention of calcium oxalate stone disease. The 3 kb Ll.LtrB intron donor cassettes from the source vector pACD4C was PCR amplified, ligated into pSIP series of lactobacillus vector pLp_3050sAmyA, yielding a novel vector pLpACD4C (8.6 kb). The quantitative real-time PCR experiment shows 94-fold increased expression of Ll.LtrB intron and 14-fold increased expression of ltrA gene in recombinant L. plantarum containing pLpACD4C. In order to target the thyA gene, the potential intron RNA binding sites in the thyA gene of L. plantarum was predicted with help of computer algorithm. The insertion location 188|189s of thyA gene (lowest E-0.134) was chosen and the wild type intron Ll.LtrB was PCR modified, yielding a retargeted intron of pLpACDthyA. The retargeted intron was expressed by using induction peptide (sppIP), subsequently the integration of intron in thyA gene was identified by PCR screening and finally ThyA- mutant of L. plantarum (ThyA18) was detected. In vitro growth curve result showed that in the absence of thymidine, colony forming units of mutant ThyA18 was decreased, whereas high thymidine concentration (10 µM) supported the growth of the culture until saturation. In conclusion, ThyA- mutant of L. plantarum (ThyA18) constructed in this study will be used as a biologically contained recombinant probiotic to deliver oxalate decarboxylase into the lumen for treatment of hyperoxaluria and calcium oxalate stone deposition.

Recombinant Lactobacillus plantarum expressing and secreting heterologous oxalate decarboxylase prevents renal calcium oxalate stone deposition in experimental rats.

BACKGROUND: Calcium oxalate (CaOx) is the major constituent of about 75% of all urinary stone and the secondary hyperoxaluria is a primary risk factor. Current treatment options for the patients with hyperoxaluria and CaOx stone diseases are limited. Oxalate degrading bacteria might have beneficial effects on urinary oxalate excretion resulting from decreased intestinal oxalate concentration and absorption. Thus, the aim of the present study is to examine the in vivo oxalate degrading ability of genetically engineered Lactobacillus plantarum (L. plantarum) that constitutively expressing and secreting heterologous oxalate decarboxylase (OxdC) for prevention of CaOx stone formation in rats. The recombinants strain of L. plantarum that constitutively secreting (WCFS1OxdC) and non-secreting (NC8OxdC) OxdC has been developed by using expression vector pSIP401. The in vivo oxalate degradation ability for this recombinants strain was carried out in a male wistar albino rats. The group I control; groups II, III, IV and V rats were fed with 5% potassium oxalate diet and 14th day onwards group II, III, IV and V were received esophageal gavage of L. plantarum WCFS1, WCFS1OxdC and NC8OxdC respectively for 2-week period. The urinary and serum biochemistry and histopathology of the kidney were carried out. The experimental data were analyzed using one-way ANOVA followed by Duncan's multiple-range test. RESULTS: Recombinants L. plantarum constitutively express and secretes the functional OxdC and could degrade the oxalate up to 70-77% under in vitro. The recombinant bacterial treated rats in groups IV and V showed significant reduction of urinary oxalate, calcium, uric acid, creatinine and serum uric acid, BUN/creatinine ratio compared to group II and III rats (P < 0.05). Oxalate levels in kidney homogenate of groups IV and V were showed significant reduction than group II and III rats (P < 0.05). Microscopic observations revealed a high score (4+) of CaOx crystal in kidneys of groups II and III, whereas no crystal in group IV and a lower score (1+) in group V. CONCLUSION: The present results indicate that artificial colonization of recombinant strain, WCFS1OxdC and NC8OxdC, capable of reduce urinary oxalate excretion and CaOx crystal deposition by increased intestinal oxalate degradation.

Genetically engineered Lactobacillus plantarum WCFS1 constitutively secreting heterologous oxalate decarboxylase and degrading oxalate under in vitro.

Hyperoxaluria is a major risk factor for urinary stone disease, where calcium oxalate (CaOx) is the most prevalent type of kidney stones. Systemic treatments of CaOx kidney stone patients are limited and comprise drawbacks including recurrence of stone formation and kidney damages. In the present work Lactobacillus plantarum (L. plantarum) was engineered to constitutively secrete oxalate decarboxylase (OxdC) for the degradation of intestinal oxalate. The homologous promoter PldhL and signal peptide Lp_0373 of L. plantarum were used for constructing recombinant vector pLdhl0373OxdC. Results showed that homologous promoter PldhL and signal peptide Lp_0373 facilitated the production, secretion, and functional expression of OxdC protein in L. plantarum. SDS-PAGE analysis revealed that 44 kDa protein OxdC was seen exceptionally in the culture supernatant of recombinant L. plantarum (WCFS1OxdC) harboring the plasmid pLdhl0373OxdC.The culture supernatant of L. plantarum WCFS1OxdC showed OxdC activity of 0.06 U/mg of protein, whereas no enzyme activity was observed in the supernatant of the wild type WCFS1 and the recombinant NC8OxdC strains. The purified recombinant OxdC from the WCFS1OxdC strain showed an activity of 19.1 U/mg protein. The recombinant L. plantarum strain secreted 25 % of OxdC protein in the supernatant. The recombinant strain degraded more than 70 % of soluble oxalate in the culture supernatant. Plasmid segregation analysis revealed that the recombinant strain lost almost 70-89 % of plasmid in 42nd and 84th generation, respectively. In conclusion, recombinant L. plantarum strain containing plasmid pLdhl0373OxdC showed constitutive secretion of bioactive OxdC and also capable of degrading externally available oxalate under in vitro conditions.

Screening of indigenous oxalate degrading lactic acid bacteria from human faeces and South Indian fermented foods: assessment of probiotic potential.

Lactic acid bacteria (LAB) have the potential to degrade intestinal oxalate and this is increasingly being studied as a promising probiotic solution to manage kidney stone disease. In this study, oxalate degrading LAB were isolated from human faeces and south Indian fermented foods, subsequently assessed for potential probiotic property in vitro and in vivo. Based on preliminary characteristics, 251 out of 673 bacterial isolates were identified as LAB. A total of 17 strains were found to degrade oxalate significantly between 40.38% and 62.90% and were subjected to acid and bile tolerance test. Among them, nine strains exhibited considerable tolerance up to pH 3.0 and at 0.3% bile. These were identified as Lactobacillus fermentum and Lactobacillus salivarius using 16S rDNA sequencing. Three strains, Lactobacillus fermentum TY5, Lactobacillus fermentum AB1, and Lactobacillus salivarius AB11, exhibited good adhesion to HT-29 cells and strong antimicrobial activity. They also conferred resistance to kanamycin, rifampicin, and ampicillin, but were sensitive to chloramphenicol and erythromycin. The faecal recovery rate of these strains was observed as 15.16% (TY5), 6.71% (AB1), and 9.3% (AB11) which indicates the colonization ability. In conclusion, three efficient oxalate degrading LAB were identified and their safety assessments suggest that they may serve as good probiotic candidates for preventing hyperoxaluria.

Synergistic Blends of Sodium Alginate and Pectin Biopolymer Hosts as Conducting Electrolytes for Electrochemical Applications.

The world needs sustainable energy resources with affordable, economic, and accountable sources. Consequently, energy innovation technologies are evolving toward electrochemical applications like batteries, supercapacitors, etc. The current study involves the solid blend biopolymer electrolyte (SBBE) with different compositions of sodium alginate blended with pectin via the casting technique. The characterization of the sample was tested by X-ray diffraction (XRD), Fourier transform infrared (FTIR) spectroscopy, AC impedance, linear sweep voltammetry (LSV), and cyclic voltammetry (CV) analyses. Evidently, the sample NP4 (NaAlg/pectin = 60:40 wt %) has a higher conductivity of 1.26 × 10-7 and 3.25 × 10-6 S cm-1 at 303 and 353 K, respectively. The performances of the samples were analyzed with variations in temperature, frequency, and time responses to signify the blended nature of the electrolyte. Hence, the studied biopolymers can be constructed for electrochemical device applications.

Evidence of insulin-dependent signalling mechanisms produced by Citrus sinensis (L.) Osbeck fruit peel in an insulin resistant diabetic animal model.

Citrus sinensis (L.) Osbeck is extensively cultivated worldwide and one of the most consumed fruits in the world. We evaluated the therapeutic properties of the methanol extract from Citrus sinensis fruit peel (CSMe) in high-fat diet-fed streptozotocin-induced insulin-resistant diabetic rats. Body weight, food intake, and water consumption were analysed. Biochemical and molecular biologic indices, and the expression of insulin receptor-induced signalling molecules were assessed to identify possible mechanisms. In addition, we conducted histology of pancreatic and adipose tissues. UHPLC-MS/MS analysis showed the presence of 17 dietary phenolics at substantial concentrations. High-fat diet-fed streptozotocin-induced diabetic rats administered CSMe (50 and 100 mg/kg) had reduced fasting blood glucose (56.1% and 55.7%, respectively) and plasma insulin levels (22.9% and 32.7%, respectively) compared with untreated diabetic control rats. CSMe reversed the biochemical abnormalities in diabetic rats, showed cytoprotective activity, and increased the intensity of the positive immunoreactions for insulin in pancreatic islets. CSMe treatment increased the expression of PPARγ in the adipose tissue and signalling molecules GLUT4 and insulin receptor. Our data suggest that CSMe could optimize glucose uptake of adipose tissues through the insulin-dependent signalling cascade mechanism and it should be investigated in the management of individuals with type 2 diabetes mellitus.

Molecular analysis of oxalate-induced endoplasmic reticulum stress mediated apoptosis in the pathogenesis of kidney stone disease.

Oxalate, a non-essential end product of metabolism, causes hyperoxaluria and eventually calcium oxalate (CaOx) stone disease. Kidney cells exposed to oxalate stress results in generation of reactive oxygen species (ROS) and progression of stone formation. Perturbations in endoplasmic reticulum (ER) result in accumulation of misfolded proteins and Ca2+ ions homeostasis imbalance and serve as a common pathway for various diseases, including kidney disorders. ER stress induces up-regulation of pro-survival protein glucose-regulated protein 78 (GRP78) and pro-apoptotic signaling protein C/EBP homologous protein (CHOP). Since the association of oxalate toxicity and ER stress on renal cell damage is uncertain, the present study is an attempt to elucidate the interaction of GRP78 with oxalate by computational analysis and study the role of ER stress in oxalate-mediated apoptosis in vitro and in vivo. Molecular docking results showed that GRP78-oxalate/CaOx interaction takes place. Oxalate stress significantly up-regulated expression of ER stress markers GRP78 and CHOP both in vitro and in vivo. Exposure of oxalate increased ROS generation and altered antioxidant enzyme activities. N-Acetyl cysteine treatment significantly ameliorated oxalate-mediated oxidative stress and moderately attenuated ER stress marker expression. The result indicates oxalate toxicity initiated oxidative stress-induced ER stress and also activating ER stress mediated apoptosis directly. In addition, the up-regulation of transforming growth factor ß-1 revealed oxalate may induce kidney fibrosis through ER stress-mediated mechanisms. The present study provide insights into the pathogenic role of oxidative and ER stress by oxalate exposure in the formation of calcium oxalate stone.

Secretion of biologically active heterologous oxalate decarboxylase (OxdC) in Lactobacillus plantarum WCFS1 using homologous signal peptides.

Current treatment options for patients with hyperoxaluria and calcium oxalate stone diseases are limited and do not always lead to sufficient reduction in urinary oxalate excretion. Oxalate degrading bacteria have been suggested for degrading intestinal oxalate for the prevention of calcium oxalate stone. Here, we reported a recombinant Lactobacillus plantarum WCFS1 (L. plantarum) secreting heterologous oxalate decarboxylase (OxdC) that may provide possible therapeutic approach by degrading intestinal oxalate. The results showed secretion and functional expression of OxdC protein in L. plantarum driven by signal peptides Lp_0373 and Lp_3050. Supernatant of the recombinant strain containing pLp_0373sOxdC and pLp_3050sOxdC showed OxdC activity of 0.05 U/mg and 0.02 U/mg protein, while the purified OxdC from the supernatant showed specific activity of 18.3 U/mg and 17.5 U/mg protein, respectively. The concentration of OxdC protein in the supernatant was 8-12 µg/mL. The recombinant strain showed up to 50% oxalate reduction in medium containing 10 mM oxalate. In conclusion, the recombinant L. plantarum harboring pLp_0373sOxdC and pLp_3050sOxdC can express and secrete functional OxdC and degrade oxalate up to 50% and 30%, respectively.

Antihyperglycemic activity and antidiabetic effect of methyl caffeate isolated from Solanum torvum Swartz. fruit in streptozotocin induced diabetic rats.

Natural remedies from medicinal plants are considered to be effective and safe alternatives to treat diabetes mellitus. Solanum torvum Swartz. fruit is widely used in the traditional system of medicine to treat diabetes. In the present study methyl caffeate, isolated from S. torvum fruit, was screened for its efficacy in controlling diabetes in animal models. Antihyperglycemic effect of methyl caffeate was studied in normal glucose-fed rats. The effects of oral administration of methyl caffeate (10, 20 and 40 mg/kg) for 28 days on body weight, fasting blood glucose, plasma insulin, hemoglobin, glycated hemoglobin, total protein, hepatic glycogen and carbohydrate metabolism enzymes in streptozotocin induced diabetic rats were investigated. Histological observations in the pancreas and GLUT4 expression in skeletal muscles were also studied. Methyl caffeate at 40 mg/kg significantly prevented the increase in blood glucose level after glucose administration at 60 min in comparison to the hyperglycemic control group. In streptozotocin induced diabetic rats, methyl caffeate produced significant reduction in blood glucose and increased body weight. The levels and/or activities of other biochemical parameters were near normal due to treatment with methyl caffeate. Methyl caffeate treated diabetic rats showed upregulation of GLUT4 and regeneration of ß-cells in the pancreas. These results substantiated that methyl caffeate possessed hypoglycemic effect, and it could be developed into a potent oral antidiabetic drug.