Type-2 diabetes is mediated by defects in either insulin secretion or insulin action. In an effort to identify extracts that may stimulate glucose uptake, similar to insulin, a high throughput-screening assay for measuring glucose uptake in skeletal muscle cells was established. During the screening studies to discover novel antidiabetic compounds from microbial resources a Streptomyces strain PM0324667 (MTCC 5543, the Strain accession number at Institute of Microbial Technology, Chandigarh, India), an isolate from arid soil was identified which expressed a secondary metabolite that induced glucose uptake in L6 skeletal muscle cells. By employing bioactivity guided fractionation techniques, a tri-substituted simple aromatic compound with anti-diabetic potential was isolated. It was characterized based on MS and 2D NMR spectral data and identified as NFAT-133 which is a known immunosuppressive agent that inhibits NFAT-dependent transcription in vitro. Our investigations revealed the antidiabetic potential of NFAT-133. The compound induced glucose uptake in differentiated L6 myotubes with an EC50 of 6.3 ± 1.8 μM without activating the peroxisome proliferator-activated receptor-γ. Further, NFAT-133 was also efficacious in vivo in diabetic animals and reduced systemic glucose levels. Thus it is a potential lead compound which can be considered for development as a therapeutic for the treatment of type-2 diabetes. We have reported herewith the isolation of the producer microbe, fermentation, purification, in vitro, and in vivo antidiabetic activity of the compound.
Viscous nature of the fermentation broth has phenomenal influence on process conditions and parameters in a fermentor. Though broth rheology has attracted significant influence in process research, still there is a challenge to modify fluid dynamics of fermentation broth. During the production of coenzymeQ 10 (CoQ 10 ) by Agrobacterium tumefaciens ATCC 4452, the culture broth becomes highly viscous due to excessive synthesis of exopolysaccharides. This hinders the CoQ 10 yield and complicates the downstream process. The present study describes how this problem was tackled by media modification and mutation. Induced mutants were generated using UV and EMS as mutagenic agents followed by rational selection based on antibiotic resistance. On screening of these mutants in sucrose based PM-2 medium, UV induced, vancomycin resistant mutant M-6, showed significant reduction (6.29 fold) in viscosity development in the broth. Mutant M-6(S), a natural variant of mutant M-6, resistant to high substrate concentration was further selected for the CoQ 10 production. Cane molasses as carbon source was found to be best suitable for CoQ 10 fermentation using mutant M-6(S). Replacing sucrose with cheaper cane molasses significantly reduced the broth viscosity with improved specific CoQ 10 content, thereby generating cost effective fermentation process. The newly developed mutant strain produced 48.89 mg/L of CoQ 10 with specific CoQ 10 content of 1.87 mg/g of DCW at 25°C, 500 rpm agitation and 0.2 vvm aeration using continuous fed batch fermentation and newly formulated cane molasses medium.
Coenzyme Q 10 (CoQ 10 ), an important antioxidant molecule playing a major role in electron transport chain, has been commercially produced by fermentation process for the use in oral nutraceutical formulations. Constructing the high-yielding CoQ 10 producing strains is a pre-requisite for cost-effective production. A superior mutant strain P-87 generated from Paracoccus denitrificans ATCC 19367, which showed 1.25-fold improvement in specific CoQ 10 content higher than the wild type strain at shake flask level, was selected to carry out the studies on CoQ 10 yield improvement through fermenter process optimization. In the course of study, initially the cane-molasses-based medium and fed-batch fermentation strategy using pHBA in combination with sucrose were standardized in shake flask using wild type strain. This strategy was subsequently translated at 2 L laboratory fermenter while optimizing the fermentation process parameters using improved mutant strain P-87. Under optimized fermentation condition, mutant strain P-87 produced 49.85 mg/L of CoQ10 having specific content of 1.63 mg/g of DCW, which was 1.36 folds higher than the specific CoQ 10 content of wild-type strain under similar optimized condition. The temperature and DO were found to be critical parameters for CoQ 10 production by mutant strain P-87. The optimum temperature was found to be 32˚C and the optimum DO concentration to be maintained throughout the fermentation cycle was found to be 30% of air saturation. Overall, a new cost-effective process has been established for the production of CoQ 10 using the cheaper substrate "cane molasses" and higher CoQ 10 producing mutant strain P-87.
*Corresponding author.
P. Tokdar et al.967
Background: Covid-19 curve can be flattened by adopting mass screening protocols with aggressive testing and isolating infected populations. The current approach largely depends on RT-PCR/rapid antigen tests that require expert personnel resulting in higher costs and reduced testing frequency. Loss of smell is reported as a major symptom of Covid-19, however, a precise olfactory testing tool to identify Covid-19 patient is still lacking.
Methods: To quantitatively check for the loss of smell, we developed an odor strip, COVID-Anosmia checker, spotted with gradients of coffee and lemon grass oil. We validated its efficiency in healthy and COVID-19 positive subjects. A trial screening to identify SARS-CoV-2 infected persons was also carried out to check the sensitivity and specificity of our screening tool.
Results: It was observed that COVID positive participants were hyposmic instead of being anosmic when they were subjected to smelling higher odor concentration. Our tool identified 97% of symptomatic and 94% of asymptomatic COVID-19 positive subjects after excluding most confounding factors like concurrent chronic sinusitis. Further, it was possible to reliably predict COVID-19 infection by calculating a loss of smell score with 100% specificity. We coupled this tool with a mobile application, which takes the input response from the user, and can readily categorize the user in the appropriate risk groups.
Conclusion: Loss of smell can be used as a reliable marker for screening for Covid-19. Our tool can rapidly quantitate anosmia, hyposmia, parosmia, and can be used as a first-line screening tool to trace out Covid-19 infection effectively.
The reduction of (RS)-2-(4-methoxyphenyl)-l, 5-benzothiazepin-3,4(2H,5H)-dione[(RS)-1] to (2S,3S)-2,3-dihydro-3-hydroxy-2-(4-methoxyphenyl)-l,5-benzothiaepin-4(5H)-one using bakers' yeast was carried out in the laboratory scale 10L fermenter. Under optimum reaction conditions, 200g of substrate was reduced to desired chiral intermediate in 4L reaction volume with 79.33% of conversion within 89h. The optimum conditions for the effective reduction were at pH 6.5 and at 30°C temperature. The crystals of desired isomer were isolated from the reaction mixture with the total yield of 42%, having 95.42% of assay purity with 99.80% of enantiomeric excess.
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