Redirecting Metabolic Flux towards the Mevalonate Pathway for Enhanced β-Carotene Production in M. circinelloides CBS 277.49

Carotenoids produced by microbial sources are of industrial and medicinal importance due to their antioxidant and anticancer properties. In the current study, optimization of β-carotene production in M. circinelloides strain 277.49 was achieved using response surface methodology (RSM). Cerulenin and ketoconazole were used to inhibit fatty acids and the sterol biosynthesis pathway, respectively, in order to enhance β-carotene production by diverting metabolic pool towards the mevalonate pathway. All three variables used in screening experiments were found to be significant for the production of β-carotene. The synergistic effect of the C/N ratio, cerulenin, and ketoconazole was further evaluated and optimized for superior β-carotene production using central composite design of RSM. Our results found that the synergistic combination of C/N ratios, cerulenin, and ketoconazole at different concentrations affected the β-carotene productions significantly. The optimal production medium (std. order 11) composed of C/N 25, 10 μg/mL cerulenin, and 150 mg/L ketoconazole, producing maximum β-carotene of 4.26 mg/L (0.43 mg/g) which was 157% greater in comparison to unoptimized medium (1.68 mg/L, 0.17 mg/g). So, it was concluded that metabolic flux had been successfully redirected towards the mevalonate pathway for enhanced β-carotene production in CBS 277.49.

Changes of platelets’ function in preeclampsia

Increased aggregation of platelets during preeclampsia was shown in several studies, yet several others reported no change. The aim of our study was to investigate platelet aggregation in a group of patients suffering from preeclampsia. In a cross-sectional study blood samples were taken from 89 hospitalized patients in the third trimester of pregnancy: 38 were suffering from mild to moderate preeclampsia and 51 patients were without preeclampsia. From the blood samples platelet aggregation, secretion of adenine nucleotides from platelets, concentration of energy-rich adenine compounds and levels of cyclic adenosine-mono-phosphate and cyclic guanosine mono-phosphate in platelets were measured. In the patients with preeclampsia, the adenosine diphosphate threshold for biphasic aggregation [odds ratio (OR):.75; 95% Confidence Interval (CI): 0.55–1.02; p<0.05], total adenine nucleotides concentration in the metabolic pool of platelets (OR:0.99; CI: 0.62–1.57; p<0.01) and cyclic adenosine-mono-phosphate (OR:0.81; CI: 0.57, 1.14; p<0.05) and cyclic guanosine mono-phosphate (OR:.78; CI: 0.55–1.09; p<0.05) levels in platelets were decreased in comparison with the control group, while adenylate energy charge in the metabolic pool of platelets (OR: >100.00; CI: 0.00->100.00; p<0.05) and secretion of adenosine triphosphate (OR:.13; CI: 0.00–14.26; p<0.05) and adenosine diphosphate (OR:.77; CI: 0.08–36.79; p<0.05) were increased. The results of our study show increased activation and aggregation of platelets in pregnant females with preeclampsia.

A Phenol Glucoside, Uncommon Coumarins and Flavonoids from Pelargonium sidoides DC

Structural examination of the metabolic pool of the aerial parts of Pelargonium sidoides DC has led to the isolation of two new metabolites, 4-allyl-2,5-dimethoxyphenol-1-β -D-glucopyranoside and 6,7-dihydroxycoumarin-8-sulfate, along with a series of uncommon compounds including (2R,3R)-(+)-dihydroquercetin-3-β -glucopyranoside, (2R,3R)-(+)-dihydrokaempferol-3-β - glucopyranoside, fraxetin-7-β -glucopyranoside, 7-methoxycoumarin-6-β -glucopyranoside, umckalin, orientin-2”-gallate, and isoorientin-2”-gallate. They are accompanied by the widespread isoorientin, orientin, fraxetin, (2R,3S)-epigallocatechin-3-gallate, gallic acid and protocatechuic acid. The structures of the compounds were established from spectroscopic studies. Determination of configurations was achieved by circular dichroism.

Chylomicron metabolism by the isolated perfused mouse heart

The catabolism of rat chylomicrons, labeled in their triacylglycerol (TG) component, was investigated using perfused working mouse hearts. Perfusion of mouse hearts with heparin increased lipoprotein lipase (LPL) activity in the perfusate. This heparin-releasable LPL pool remained constant over a variety of experimental conditions, including workload and fatty acid concentrations, making the mouse heart a suitable model to study chylomicron catabolism. Endothelium-bound LPL hydrolyzed radiolabeled 3H-labeled chylomicrons (0.4 mM TG); the fate of LPL-derived 3H-labeled fatty acids was split evenly between oxidation (production of3H2O) and esterification (incorporation into tissue lipids, mainly TG). In comparison, the oxidation of 0.4 mM [3H]palmitate complexed to albumin was fourfold greater than esterification into tissue lipids. Surprisingly, the addition of unlabeled palmitate (0.4 or 1.2 mM) to perfusions with3H-chylomicrons did not affect the fate (either oxidation or esterification) of LPL-derived 3H-fatty acids. These results suggest that fatty acids produced from lipoprotein hydrolysis by the action of LPL and fatty acids from a fatty acid-albumin complex do not enter a common metabolic pool in the heart.

Effects of aminooxyacetate on glutamate compartmentation and TCA cycle kinetics in rat hearts

The nonspecific transaminase inhibitor aminooxyacetate (AOA) has multiple influences on the dynamics of13C appearance in glutamate in rat hearts as measured by 13C nuclear magnetic resonance (NMR) without altering O2 consumption or tricarboxylic acid (TCA) cycle flux. These include the following: 1) a reduced rate of13C enrichment at glutamate C3 and C4; 2) a near coalescence of the C3 and C4 fractional enrichment curves; 3) a dramatic alteration in the time-dependent evolution of the glutamate C4 multiplets, C4S and C4D34; and 4) a decrease in the NMR visibility of glutamate. A fit of the13C fractional enrichment curves of glutamate C4 and C3 in the absence of inhibitor to a kinetic model of the TCA cycle gave values for transaminase flux of 7.5 μmol ⋅ min−1 ⋅ g dry wt−1 and TCA cycle flux of 7.5 μmol ⋅ min−1 ⋅ g dry wt−1, thereby confirming reports by others that the kinetics of13C enrichment of glutamate C3 and C4 in heart tissue is significantly affected by flux through reactions other than TCA cycle. The 13C fractional enrichment data collected in the presence of 0.5 mM AOA could not be fitted using this same kinetic model. However, kinetic simulations demonstrated that the time-dependent changes in C4S and C4D34 are only consistent with a 10-fold reduction in the size of intermediate pools undergoing rapid turnover in the TCA cycle. We conclude that inhibition of glutamic-oxalacetic transaminase by AOA effectively reduces the size of the α-ketoglutarate pool in rapid exchange with the TCA cycle. Our data indicate that changes in glutamate multiplet areas in the13C NMR spectra of heart (as demonstrated by glutamate C4S and C4D34) are more sensitive to alterations in metabolic pool sizes in exchange with the TCA cycle than are measurements of 13C fractional enrichment at glutamate C3 and C4.