The effect of annatto on insulin binding properties in the dog

Various plants are used in Caribbean folklore for the treatment of a variety of illnesses including diabetes mellitus. Preliminary investigations of several crude plant extracts have indicated that the annatto (Bixa orellana), among others, does in fact exhibit hypoglycaemic properties. This present investigation sought to isolate the hypoglycaemic principle(s) from the crude extract and to determine the mechanism of action. Purification experiments employing thin layer chromatography (TLC) and high performance liquid chromatography (HPLC) resulted in an oil-soluble, partially purified annatto extract. The latter caused a decrease in blood glucose level of 5.62+/-0.13 (n=34) mmol/dL versus 6.31+/-0.12 (n=34) for the control (p<0.01) 1 h after administration. This hypoglycaemia persisted for an additional hour when the oral glucose tolerance test (OGTT) was performed on dogs treated with annatto and compared with the control. Plasma insulin levels measured at 1.0 h showed that there was an increase in plasma insulin levels of 59.57+/-8.3 microIU/mL for the annatto treated dogs versus 40.95+/-5.46 microIU/mL for the control (p<0.05), this elevation persisted throughout the duration of the OGTT which followed. Insulin receptor studies, using a modification of the method of Gambhir et al. done on mononuclear leucocytes and erythrocytes obtained from blood taken 1 h after administration showed that there was an increase in the percentage receptor binding when compared with the control. Insulin affinity results showed that there was an increase of 1.8+/-0.2x10(8) m-1 (n=12) in mononuclear leucocytes for the annatto treated dogs versus 1.2+/-0.2x10(8) m-1 for the control (p<0.05). In the enythrocytes, there was also an increase in affinity from 1.2+/-0.2x10(8) m-1 to 2.3+/-0.2x10(8) m-1 for the control and treated animals, respectively. In conclusion, it can be stated that annatto is responsible for the hypoglycaemic episodes seen in the dogs which was mediated by an increase in plasma insulin concentration as well as an increase in insulin binding on the insulin receptor due to elevated affinity of the ligand for the receptor.