Potent in Vitro α-Glucosidase Inhibition of Secondary Metabolites Derived from Dryopteris cycadina.

α-glucosidase is responsible for the hydrolysis of complex carbohydrates into simple absorbable glucose and causes postprandial hyperglycemia. α-glucosidase inhibition is thus the ideal target to prevent postprandial hyperglycemia. The present study was therefore designed to analyze the effects of various compounds isolated from Dryopteris cycadina against α-glucosidase including ß-Sitosterol 1, ß-Sitosterol3-O-ß-d-glucopyranoside 2, 3, 5, 7-trihydroxy-2-(p-tolyl) chorman-4-one 3, Quercetin-3-0-ß-d-glucopyranoside (3/→0-3///)- ß-d- Quercetin -3-0- ß â»d-galactopyranoside 4 and 5, 7, 4/-Trihydroxyflavon-3-glucopyranoid 5. The in vitro spectrophotometric method was used for the analysis of test compounds against possible inhibition. Similarly, molecular docking studies were performed using the MOE software. These compounds showed concentration-dependent inhibition on α-glucosidase, and compounds 1 (IC50: 143 ± 0.47 µM), 3 (IC50:133 ± 6.90 µM) and 5 (IC50: 146 ± 1.93 µM) were more potent than the standard drug, acarbose (IC50: 290 ± 0.54 µM). Computational studies of these compounds strongly supported the in vitro studies and showed strong binding receptor sensitivity. In short, the secondary metabolites isolated from D. cycadina demonstrated potent α-glucosidase inhibition that were supported by molecular docking with a high docking score.

Biotechnology and molecular biology of the alpha-glucosidase inhibitor acarbose.

The alpha-glucosidase inhibitor acarbose, O-[4,6-dideoxy-4[1 s-(1,4,6/5)-4,5,6-trihydroxy-3-hydroxymethyl-2-cyclohexen-1-yl]-amino-alpha-D-glucopyranosyl]-(1-->4)- O-alpha-D-glucopyranosyl-(1-->4)-D-glucopyranose, is produced in large-scale fermentation by the use of strains derived from Actinoplanes sp. SE50. It has been used since 1990 in many countries in the therapy of diabetes type II, in order to enable patients to better control blood sugar contents while living with starch-containing diets. Thus, it is one of the latest successful products of bacterial secondary metabolism to be introduced into the pharmaceutical world market. Cultures of Actinoplanes sp. also produce various other acarbose-like components, of which component C is hard to separate during downstream processing, which is one of the most modern work-up processes developed to date. The physiology, genetics and enzymology of acarbose biosynthesis and metabolism in the producer have been studied to some extent, leading to the proposal of a new pathway and metabolic cycle, the "carbophore". These data could give clues for further biotechnological developments, such as the suppression of side-products, enzymological or biocombinatorial production of new metabolites and the engineering of production rates via genetic regulation in future.

Reduced formation of byproduct component C in acarbose fermentation by Actinoplanes sp. CKD485-16.

Acarbose fermentation was conducted by cultivation of Actinoplanes sp. CKD485-16. Approximately 2,300 mg/L of acarbose was produced at the end of cultivation along with 600 mg/L of the acarbose byproduct component C. Maltose, a known moiety of acarbose, should be maintained at high concentration levels in culture broths for efficient acarbose production. The acarbose yield increased with an increasing osmolality of the culture medium, with a maximum value of 3,200 mg/L obtained at 500 mOsm/kg. Component C was also produced in proportion to the osmolality. Conversion of acarbose to component C was accomplished with resting whole cells. Inhibitors of the conversion of acarbose to component C were sought since component C is probably derived from acarbose. Valienamine was found to be a potent inhibitor, resulting in a more than 90% reduction in component C formation at a 10 microM concentration. Effects were similar in a 1,500-L pilot fermentor with acarbose and component C yields of 3,490 and 43 mg/L at 500 mOsm/kg, respectively.

Synthesis of acarbose analogues by transglycosylation reactions of Leuconostoc mesenteroides B-512FMC and B-742CB dextransucrases.

Two new acarbose analogues were synthesized by the reaction of acarbose with sucrose and dextransucrases from Leuconostoc mesenteroides B-512FMC and B-742CB. The major products for each reaction were subjected to yeast fermentation, and then separated and purified by Bio-Gel P2 gel permeation chromatography and descending paper chromatography. The structures of the products were determined by one- and two-dimensional 1H and 13C NMR spectroscopy and by matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS). B-512FMC-dextransucrase produced one major acarbose product, 2(I)-alpha-D-glucopyranosylacarbose and B-742CB-dextransucrase produced two major acarbose products, 2(I)-alpha-D-glucopyranosylacarbose and 3(IV)-alpha-D-glucopyranosylacarbose.

Addition of maltodextrins to the nonreducing-end of acarbose by reaction of acarbose with cyclomaltohexaose and cyclomaltodextrin glucanyltransferase.

New kinds of acarbose analogues were synthesized by the reaction of acarbose with cyclomaltohexaose and cyclomaltodextrin glucanyltransferase (CGTase). Three major CGTase coupling products were separated and purified by Bio-Gel P2 gel-permeation chromatography. Digestion of the three products by beta-amylase and glucoamylase showed that they were composed of maltohexaose (G6), maltododecaose (G12), and maltooctadecaose (G18), respectively, attached to the nonreducing-end of acarbose. 13C NMR of the glucoamylase product (D-glucopyranosyl-acarbose) showed that the D-glucose moiety was attached alpha- to the C-4-OH group of the nonreducing-end cyclohexene ring of acarbose, indicating that the maltodextrins were attached alpha-(1-->4) to the nonreducing-end cyclohexene of acarbose.