Identification and Quantification of Aldose Reductase Inhibitory Flavonoids in Herbal Formulation and Extract of Gymnema sylvestre Using HPLC-PDA and LC-MS/MS

Adulteration of herbal supplements is a major issue for many countries. A simple and reliable HPLC-PDA method was developed for quantification of aldose reductase inhibitory flavonoids rutin, quercetin, and kaempferol. The chromatographic separation was performed on a Fortis C18 column in gradient mode with detection at 267？nm. The presence of these markers was confirmed through the accurate m/z values and MS/MS data obtained using quadruple time of flight mass spectrometry (QTOF-MS). The proposed method was successfully applied to examine the amount of these active constituents in antiobese polyherbal formulation and plant extract of Gymnema sylvestre. 1. Introduction Antiobese polyherbal formulation (Ayurslim) is usually used in ayurvedic medicine to treat various obesity-related disorders like diabetes, hypertension, atherosclerosis, gall bladder problems, and certain types of cancer. It comprises five ingredients such as Garcinia cambogia, Gymnema sylvestre, Commiphora wightii, Terminalia chebula, and Trigonella foenum-greacum. These ingredients are present in ayurslim and have considerable effect on weight reduction and lipid profile as it was shown in the report on evaluation of clinical efficacy of ayurslim on body weight, body mass index, lipid profile, and skin fold thickness [1]. Flavonoids occupy a wide space of biologically active compounds that are omnipresent in plants and obtained a lot of significance due to their presence in the human diet, which in turn imparts some favourable effects to promote good health. Aldose reductase is an enzyme which converts the glucose into sorbitol and then fructose. The accumulation of sorbitol in the tissues causes swelling and in turn leads to secondary complications of diabetes like obesity. Earlier studies have revealed that flavonoids such as quercetin [2, 3], rutin [4], and kaempferol [5] were recommended to be responsible for antiobesity activity, followed by inhibiting aldose reductase enzyme [6]. The chemical structures and the proposed fragmentation pattern were shown in Figure 1. In addition to this, flavonoids have exhibited profound pharmacological activities like anti-inflammatory, antioxidant, antithrombotic, hepatoprotective, antiviral, and other biological activities [7]. Reports which can be cited with respect to simultaneous estimation of these three components were rather limited. Till now four HPLC reports are available on simultaneous estimation of these flavonoids in the plant extract of sea buckthorn [8], in Melia azedarach Linn. leaves [9], in the leaf extracts of Moringa oleifera