Isolation And Antihyperglycemic Effect Of Chemical Constituents From Rhizoma Coptidis

Rhizoma Coptidis is one of traditional medicinal plants, often used in treatment of clearing heat and detoxicating. Modern pharmacology studies showed it owned many activities, such as antimicrobial, anti-inflammation, anticancer, antihypertension, antihyperglycemic, antioxidant, and so forth. Its main constituents are alkaloids, shuch as berberine, coptistine, epiberberine, palmatine, jatrorrhizine. Of them, berberine had the highest mass ratio in Rhizoma Coptidis, about6%-10%. Many studies showed that berberine was main active constituent of Rhizoma Coptidis, but berberine cann't take the place of Rhizoma Coptidis, one reason was that beberine existed in many plants, another one was that the content of berberine from Rhizoma Coptidis wasn't the highest in all plants. In Chinese traditional medicinal study, Rhizoma Coptidis often was used by apozem and powder in clinic remedy, and some reports suggested that it showed some better pharmacological effect than berberine. In order to clarify material base of pharmacological effect derived from Rhizoma Coptidis, we made some study as follows:1. Study on chemical constituents from Rhizoma CoptidisTo study polar of constituents from Rhizoma Coptidis by High Speed Counter Current Chromatography (HSCCC). HSCCC is liquid-liquid distribution chromatography technology. Rhizoma Coptidis extracts was isolated in CHCl3-MeOH-0.2MHCl olvent system by HSCCC. The result showed that most of constituents from Rhizoma Coptidis were hydrophilic compounds, about50%ass ratio of alcohol extracts. Six main alkaloids were isolated in one run by HSCCC, purify of them were all above85%.Rhizoma Coptidis extracts was firstly extracted by different polar solvents, petroleum ether, chloroform, and n-butyl alcohol. Petroleum ether extracts had little. Petroleum extracts, chloroform extracts and n-butyl alcohol extracts occupied respectively5%,10%, and70%ass ratio of Rhizoma Coptidis alcohol extracts. Then slovent extracts were separated by traditional solum chromatography technology. From them,14compounds were isolated and identified by TLC, HPLC, NMR and MS. They were berberine, berbamine, palmatine, coptisine, berberubine, epiberberine, magnoflorine, groenlandicine, ferulic acid, choline, jatrorrhizine, columbamine, oxyberberine, and8-oxocoptisine. Choline was the first time isolated from coptidis plants.Rhizoma Coptidis extracts was analysised with the method of content determination in2010Chinese Pharmacopoeia (2010edition).8compounds were identified in fingerprint of HPLC. Ferulic acid, magnoflorine and groenlandicine were determined by quantitative analysis with diode array detector.2. Antihyperglycemic effect of constituents from Rhizoma CoptidisHepG2cells is similar to human liver cells. Effect of glucose metabolism resulted from different constituents was determined by the glucose consumption in HepG2cells and alloxan-induced diabetes. Almost all compounds can increase glucose consumption in HepG2cells, and had dose-dependent relationship within test concentration. In3μM centration, berberine had the strongest activity, similar to positive control metformin, secondly, coptisine and jatrorrhizine also can promote glucose consumption in cells, epiberberine, columbamine and berberubine showed similar effect to ferulic acid, choline showed little activity. Main alkaloids had interaction glucose-lowering effect on HepG2cells, so do berberine and ferulic acid or choline. Compared with same dose of berberine, berberine38mg-L-in combination with choline100mg-L"1can make glucose consumption increase by34%nd elevate cell livability up to75%n HepG2cells. Berberine and ferulic acid had obvious synergistic glucose-lowering effect on HepG2cells as their mass ratio was100:1. Total alkaloids showes better activity than Rhizoma Coptidis ethanol extracts. Constituents except for main alkaloids had more little effect. Animal experiments had same effect as cells assay.3. Antioxidant effect of constituents from Rhizoma Coptidis in vitroThe antioxidant effect of compounds from Rhizoma Coptidis was evaluated by colorimetric method on scavenging superoxide anion, hydroxyl free radical and hydrogen peroxide in vitro. ROS generated in mitochondria and HepG2cells was determined by fluorescence method. The result suggested almost compouds had antioxidant activities within test concentration. As a whole, berberubine, groenlandicine and coptisine has higher activities than berberine.