Advancement of prodrug approaches for nucleotide antiviral agents

: Synthetic nucleoside or nucleotide analogues played a key role to the development of antiviral agents in past decades. However, low membrane permeability and insufficient cellular phosphorylation impaired the biological activity of polar nucleoside drugs because they have to penetrate the cell membrane and be phosphorylated to active metabolite in stepwise by intracellular enzymes. To overcome these limitations, diverse lipophilic prodrug modifications based on nucleoside mono-, di-, and triphosphate were designed and put into practice to efficiently deliver nucleoside into the target site, and bypass the rate-limited phosphorylation step. As the most successful prodrug strategy, ProTide technology has led to the discovery of three FDA-approved antiviral agents including sofosbuvir, tenofovir alafenadmide, and remdesivir which has been authorized for emergency use in patients of COVID-19 in the US. In recent years, nucleoside di- and triphosphate prodrugs have also made the significant progress. This review will focus on the summary of design approach and metabolic activation path of different nucleotide prodrug strategies. The potential application of nucleotide prodrugs for treatment of COVID-19 was also described due to the pandemic of SARS-CoV-2.

Mineralization of Melanoidin by H₂O₂ Producing Enzymes from Marine Cyanobacteria Oscillatoria boryana BDU 92181

Intracellular enzymes of Oscillatoria boryana BDU 92181 exhibited mineralizing activity on melanoidin, a recalcitrant pigment present in the distillery wastewater. Melanoidin decolourization was postulated to be due to the production of hydrogen peroxide and molecular oxygen released by the cyanobacterium during photosynthesis. The present study was aimed to find out the efficacy of the marine cyanobacterium O. boryana BDU 92181 in producing H2O2 and enzymes involved in hydrogen peroxide production with a view to utilize its potential for decolorization of melanoidin pigment in the distillery effluent. The enzymes involved in the melanoidin degradation have not so far been attempted with cyanobacteria. The results obtained in the present work suggested the activity of the glucose oxidase and Manganese peroxidase enzymes in a marine cyanobacterium Oscillatoria boryana BDU 92181 and whose activity was found to be enhanced in the presence of melanoidin.

Early diagnostic and prognostic criteria for health disorders in chemical workers

The production of synthetic materials - fiberglass and rubber-technical products, as well as products derived from oil refining wastes continues to grow. A characteristic feature of the working conditions of these enterprises is the content of chemicals of hazard class 1-4, which may be potentially hazardous to workers. It imperative to justify prenosological laboratory criteria for health problems of manufacturing workers. The results of the hematological, biochemical, immunological and cytochemical parameters among manufacturing workers are presented. Significant changes in the processes of oxidative metabolism, lipid metabolism, impaired immune status, and intracellular enzyme activity were found in a significant proportion of the workers. Changes were detected in workers even with a short work experience (5 years) and of young age (20-30 years). With an increase in the occupational length of service, the frequency of these disorders increased. Based on the results obtained, early diagnostic and prognostic criteria for health disorders in workers of diverse chemical industries are substantiated. Based on the results obtained, early diagnostic and prognostic criteria for health disorders in workers of diverse chemical industries are substantiated. For rubber manufacturing workers: increased malonic dialdehyde, reduced catalase activity, the activity of catalase is lowering; increased enzymes activity (AST, ALT, GGT), protein abnormalities, erythropenia, decreased hemoglobin levels, increased reticulocyte. For fiber glass manufacturing workers: indicators of the oxidative antioxidant system, hemograms - increased lymphocytes, erythrocytosis, leukocytosis, increased CD3 +, increased CD4 +, CD 16+, CD 20 +, FAL, IgG, increased total protein, hyperfermentemia (GGT, ALT), changes in the intracellular enzymes activity - acid phosphatase of neutrophils and lymphocytes, decreased myeloperoxidase, increased glycogen levels. For olefin oxides manufacturing workers - this is hyperfermentemia (ALT, alkaline phosphatase), protein metabolism changes, increased red blood cells, reticulocytes, neutrophils, lymphocytes, eosinophils, platelets, leucopenia, decreased FAL, increased IgM, decreased IgA, IgG, changes in the intracellular enzymes activity - decrease myeloperoxidase, increase acid phosphatase of neutrophils and lymphocytes.

Degradation and Detoxification of Aflatoxin B1 by Tea-Derived Aspergillus niger RAF106

Microbial degradation is an effective and attractive method for eliminating aflatoxin B1 (AFB1), which is severely toxic to humans and animals. In this study, Aspergillus niger RAF106 could effectively degrade AFB1 when cultivated in Sabouraud dextrose broth (SDB) with contents of AFB1 ranging from 0.1 to 4 μg/mL. Treatment with yeast extract as a nitrogen source stimulated the degradation, but treatment with NaNO3 and NaNO2 as nitrogen sources and lactose and sucrose as carbon sources suppressed the degradation. Moreover, A. niger RAF106 still degraded AFB1 at initial pH values that ranged from 4 to 10 and at cultivation temperatures that ranged from 25 to 45 °C. In addition, intracellular enzymes or proteins with excellent thermotolerance were verified as being able to degrade AFB1 into metabolites with low or no mutagenicity. Furthermore, genomic sequence analysis indicated that the fungus was considered to be safe owing to the absence of virulence genes and the gene clusters for the synthesis of mycotoxins. These results indicate that A. niger RAF106 and its intracellular enzymes or proteins have a promising potential to be applied commercially in the processing and industry of food and feed to detoxify AFB1.

Freeze-thaw system for thermostable β-Galactosidase isolation from Gedong Songo Geobacillus sp. isolate

The effective isolation of intracellular enzymes from thermophilic bacteria is challenging because of their sturdy membrane. On the other hand, the low-cost and nontoxic method is essential for industrial food enzymes. The freeze-thaw cycles using acetone-dry ice as a frozen system was studied for efficient isolation of thermostable b-galactosidase from Geobacillus sp. dYTae-14. This enzyme has been known for application in the dairy industry to reduce the lactose content. In this study, the freeze-thaw method was performed with cycle variations 3, 5, and 7 cycles. Acetone-dry ice (-78°C) is used as a frozen system and boiling water for thawing. The b-galactosidase activity was assayed using ortho-Nitrophenyl-β-galactoside (ONPG) as substrate and protein content determined with the Lowry method. The results show that the most effective freeze-thaw is five cycles. The enzyme’s highest specific activity is 3610.13 units/mg proteins at 40-60 % ammonium sulfate saturation, with a purity value of 2.52.

A Review of Microvascular Complications

Complications of Diabetes mellitus include chronic damage, dysfunction and disappointment of various organs mainly affecting the eyes, kidneys, heart and blood vessels. Various minerals and vitamins acts as cofactors in regulation of insulin secretion and action ,among which magnesium dramas a key role. Magnesium is an essential mineral and plays a major role in carbohydrate metabolism. It acts as a co-factor in glucose transport mechanism and for various intracellular enzymes involved in carbohydrate oxidation. Deficiency of magnesium results in increase in insulin resistance and decrease in glucose uptake of the cells in the body. Serum magnesium levels in healthy individuals are constant, but 25 to 39% people with Type 2DM have low levels of magnesium. Hypomagnesemia has been found to have deleterious effect on glucose homeostasis and insulin sensitivity in Type 2DM. Low levels of magnesium have also been attributed to the progression and development of micro and macrovascular difficulties in Type 2DM.

Analysis of biochemical markers in the saliva and correlation with clinical parameters in patients with aggressive periodontitis, before and after the therapy

Introduction/Objective. Aggressive periodontitis (AP) is a progressive disease that leads damage to periodontal tissues. The aim of the study was the analysis of intracellular enzymes: aspartate aminotransferase (AST), alanine aminotransferase (ALT), alkaline phosphatase (ALP), acid phosphatase (ACP) and electrolytes in the saliva of patients with AP and their correlation with clinical parameters before and after the therapy. Methods. The study included 30 patients with AP (experimental group) and 35 patients with healthy parodontium (control group). Intracellular enzymes and electrolytes were analyzed in an unstimulated saliva of subjects with AP, before and after the therapy and in saliva of the control group. The analysis of biochemical markers was carried out using kinetic methods with commercial reagents. Results. Concentrations of the biochemical markers AST (28.18 ? 25.16), ALT (5.48 ? 5.14), ALP (31.13 ? 37.79), ACP (17.53 ? 14.77), calcium (2.80 ? 1.97), phosphate (4.43 ? 1.92) in the saliva of subjects of the experimental group were statistically significantly higher in relation to the control group (p = 0.000; p = 0.001). Significant correlation was found between AST values, debris index (? = -0.444; p = 0.026) and calculus index (? = -0.513; p = 0.009), and between the plaque index and ALP level in the saliva after therapy (p = 0.020). Conclusion. The investigation will contribute to a better understanding and standardization of biomarkers in the saliva that mayhelp in diagnosing the AP and evaluation of the applied therapy.

Fatty Acid Storage from Systemic FFA and VLDL-triglyceride Between Visceral and Subcutaneous Adipose Tissue Depots in Morbid Obesity (OR09-06-19)

Objectives Understanding both systemic and cellular factors that govern lipid storage in different tissue depots may be essential for understanding the genesis of differences in body fat distribution. Methods We assessed fatty acid (FA) storage in upper body subcutaneous adipose (UBSQ), visceral (omental) adipose (VAT), and liver from systemic free fatty acids (FFA) using [9,10–3H] palmitate tracer and from very low density lipoprotein (VLDL) triglycerides (TG) using autologous [1–14C]VLDL-TG tracers combined with carefully timed biopsies in patients undergoing bariatric surgery. We measured the activity of 4 enzymes that regulate adipocyte FA storage: lipoprotein lipases (LPL) – an extracellular enzyme, and 3 intracellular enzymes - acyl-CoA synthase (ACS), glycerol-3-phospate acyltransferase (GPAT) and diacylglycerol transferase (DGAT). Finally, we examined systemic factors (fasting plasma insulin and resting energy expenditure (REE)) that might relate to lipid storage. Results Storage of systemic FFA was well correlated between VAT and UBSQ (r = 0.53, P < 0.006), UBSQ and liver (r = 0.69, P = 0.0003), but not between VAT and liver. Storage of VLDL-TG was positively correlated between all three depots. Fasting LPL activity was positively correlated with storage of VLDL-TG in VAT (r = 0.47, P < 0.05) but not UBSQ. Fasting palmitate was correlated with storage rates of systemic FFA in both VAT (r = 0.45, P < 0.03) and UBSQ (r = 0.48, P < 0.03). The activities of intracellular storage enzymes (ACS, DGAT, and GPAT) were unrelated to UBSQ storage of systemic FFA or VLDL-TG. None of the intracellular enzymes correlated with storage of systemic FFA in VAT; only ACS (r = 0.50, P < 0.03) was positively correlated with VLDL-TG storage; DGAT and GPAT tended to correlate (P = 0.056 and P = 0.0719, respectively). Plasma insulin was unrelated to storage of either lipid in both depots. Interestingly, REE was negatively correlated to VAT storage of VLDL-TG (r = 0.45, P < 0.03) but not systemic FFA, whereas REE was negatively correlated to UBSQ storage of systemic FFA (r = 0.50, P < 0.05) but not VLDL-TG. Conclusions In morbid obesity, differences in cellular and systemic factors contribute to variations in storage of VLDL-TG versus systemic FFA between VAT and UBSQ; however, within each subject there is no preferential partitioning of FA to either VAT or UBSQ. Funding Sources DK40484, DK45343, DK50456.