Analysis Of Metabolic Components In Rats And Its Inhibitory Activity Against Aldose Reductase

Objective:Radix Scrophulariae is dried roots of the Scrophularia ningpoensis Hemsl. It is widely used in clinical therapy due to its function of purging fire, nourishing yin, cooling blood and detoxification. As a kind of pure traditional Chinese medicine, Fufang Xueshuantong capsule has been the initial medicine for the patients with diabetes in Chinese-Western-Combined Therapy. However, the contribution of Radix Scrophulariae to the efficacy of capsule is not clear. Modern pharmacological studies have pointed out that Radix Scrophulariae had the effect of reducing blood sugar, enhancing immunity, anti-oxidation and anti-thrombosis. Consulting to past 10 years of Chinese and English datum, which concentrated on the work of the Radix Scrophulariae pharmaceutical researches, we have found that they focused mainly on the quality control and pharmacological researches. There are less reports on the the material foundation and process in vivo of Radix Scrophulariae at present. In order to elucidate bioactive constituents of Radix Scrophulariae in Xueshuantong capsule, the chemical constituents of the ethanol extract of Radix Scrophulariae, and metabolism in rats were initially studied. Secondly, the research studied inhibition of components absorbed into plasma on aldose reductase.Methods:1. We systematically refered to the Chinese and foreign literature, and sorted the literature on chemical composition, content determination, pharmacology and toxicology, effective substance and clinical research. This could lay a solid foundation for post-experimental study.2. According to the production technology, we have got reference dosage and extraction method. The mobile phase consisted of acetonitrile and water(with 0.1% formic acid) was used as gradient elution for separation. Samples were analyzed by a reversed-phase C18 column using HPLC-MS techniques with an ESI source. We identified the chemical components of the ethanol extract of Radix Scrophulariae, and analyzed the metabolic components in rat plasma, urine and feces after oral administration of Scrophularia ningpoensis extract.3. Referring to our earlier research, the subject studied the inhibition of prototype compounds on aldose reductase.Results:1. The results showed that in MS detection, the main chemical components in the ethanol extract of Radix Scrophulariae were easily to become negative ions in the electrospray ionization procedure and they had srong responses.37 compounds from Scrophularia ningpoensis extract were detected including 12 iridoid glycosides,20 phenylpropanoids and 5 unknown compounds. Thus, the characteristic groups of iridoid glycosides and phenylpropanoids were determined preliminarily. This method has an advantage of simple operation, rapid measurement. It is a powerful tool for identification of chemical constituents of the extract of Radix Scrophulariae.2. After oral administration of extract of Radix Scrophulariae, the rat’s plasma, urine and feces were also analyzed. In vivo, it was comfirmed that harpagide, harpagoside and angoroside C could enter into the blood in prototype forms. Besides, another 2 prototype compounds and 2 metabolites were detected in rat plasma. A total of 92 chemical constituents were detected in urine and feces consequently, concluding 34 prototypes,31 metabolites and 27 unknown compounds.3. From the perspectives of metabolites in rat plasma and its identification in vitro activity, we selected the main four prototype components from the constituents absorbed into plasma: harpagide, harpagoside, angoroside C and cinnamic acid. The inhibition of prototype compounds on AR were studied. The results showed that cinnamic acid had a smaller impact on the inhibition rate, while harpagide, harpagoside and angoroside C did not exhibit potent activity.Conclusion:Our study established methods for identifying the chemical constituents of the extract of Radix Scrophulariae and metabolites in vivo by HPLC-ESI-MS. Take the chemical ingredients profile, metabolite profile in vivo and vitro activity of Radix Scrophulariae as a main line, the research provides scientific basis for pharmacodynamic constituents and action mechanism of Radix Scrophulariae and Fufang Xueshuantong capsule, especially in the treatment of diabetic retinopathy.