INHIBITION OF HUMAN HAEMOGLOBIN GLYCOSYLATION BY FLAVONOIDS CONTAINING METHANOLIC EXTRACTS OF THE LEAF OF Cnestis ferruginea

Abstract

Protein glycosylation by a sugar adduction, an oxidative process is exemplified by the glycosylation of haemoglobin A. The objective of this study, therefore, is to assess the influence of the leaf extracts of Cnestis ferruginea on human haemoglobin glycosylation. The methanolic extracts of the leaf of cnestis ferruginea were prepared and tested for the presence of flavonoids and the total phenolic content obtained was 0.0175mg/ml Catechin. Haemoglobin was partially purified from the erythrocytes of normal and diabetic individuals according to the method of Asgary et al. (1999). An assessment of the effect of glucose on the time-dependent glycosylation of haemoglobin shows that the degree of human haemoglobin glycosylation increased with the period of incubation in a concentration-dependent manner up to 20mg glucose/ml. A study of the effect of varying concentrations (10-30ug/ml) of the flavonoid-containing methanolic extracts of Cnestis ferruginea (Cn,f) and quercetin (Q), a known antioxidant, reduced the extent of haemoglobin glycosylation at all the concentrations used irrespective of the incubation period. Glycosylation in the presence of 20mg glucose/ml was about 72% and varying concentrations (10, 20, 30ug/m ) of Cn.f reduced the glycosylation to 41, 31, and 29% respectively. Similar effects were obtained for quercetin. The observed inhibition on glycosylation was concentration and time dependent. Results obtained from the investigation carried out on the possible effect of the extracts of Cn.f on haemoglobin glycosylation in the present of physiological concentrations of glucose ( ≤1mg/ml) indicated that there was no significant glycosylation (P≥0.05) at the varied concentration of glucose up to 1mg/ml, 4% inhibition of glycosylation was observed after 24hr in the presence of 10ug/ml extract of Cn.f. The extent of inhibition increased from 5.5 to 7% after 48 and 72hr respectively. Using 20 and 30µg/ml methanolic extracts of Cn.f and quercetin, the inhibition was total even after prolonged hours of incubation. Statistical analysis of these results showed that there was no significant difference (P>0.05) in the inhibitory effects of methanolic extracts of cn.f and quercetin on haemoglobin glycosylation at the concentrations of glucose used in this study. On the contrary, quercetin and α-tocopherol, a naturally occurring antioxidant, which has been widely used as dietary supplement, and extracts of Cn.f did not show any significant effect ( ≤14%) on glycosylation haemoglobin (HbA) in diabetic subjects. Although, the mechanism of inhibition of haemoglobin glycosylation is not yet clear, it seems likely that quercetin and leaf extracts of Cn.f being very rich in flavonoids could be converted into glycoside adducts for there to be a reduction in haemoglobin glycosylation. In view of the fact that methanolic extracts of cn.f inhibited glycosylation, leaves of cnestis ferruginea may be used as food supplement in order to prevent or inhibit haemoglobin glycosylation diabetics. In conclusion, leaf extract of cnestis ferruginea inhibited haemoglobin glycosylation possibly by preventing the condensation of glucose with the N-terminal amino group of β-chains of HbA.