| Malus hupehensis(Pamp.)Rehd.commonly named as Hupeh crabapple,Chinese crabapple or tea crabapple,is a Chinese native species of flowering plant in the apple genus Malus of the family Rosaceae.It prevalently distributes in the south of China such as Hubei,Sichuan and Jiangxi provinces,etc.Its leaves have been locally used as a kind of Non-Cuamellia-Tea for more than 400 years,especially as commomly used herbal tea and herbal medicine in Tujia ethnic minority in Huibei Province of China.Modern pharmacological studies have shown that Hupeh crabapple possesses various biological activities such as hypoglycemic,hypolipidemic,antioxidative,estrogen-like effects and anti-pathogenic microorganisms.In this study,we investigate the chemical constituents of Hupeh crabapple collected in Hubei Province and develop a rapid and validated method for its quality assessment to provide scientific evidences for further utilization Hupeh crabapple.Sixteen compounds were isolated from the 70%ethanolic extractives of the leaves of Hupeh crabapple by using a variety of column chromatographies of silica gel,Sephadex LH-20 gel and preparative HPLC.Their chemical structures were elucidated as chlorogenic acid(1),protocatechuic acid(2),cryptochlorogenic acid(3),neo-chlorogenic acid(4),caffeic acid(5),P-hydroxy-phenylpropionic acid(6),3-O-coumaroylquinic acid(7),cis-coumaric acid(8),phloroglucinol(9),(β-hydroxypropiovanillone(10),3-hydroxy-1-(4-hydro xy-3,5-dimethoxyphenyl)-1-propanone(11),3-((P)-2’,4,4’,5-tetrahydroxy-6’-(β-D-glucopyranosyloxy)-[1,1’-biphenyl]-2-yl)propanoicacid(12),3-((M)-2’,4,4’,5-tetrahydroxy-6’-(β-D-glucopyranosyloxy)-[1,1’-biphenyl]-2-yl)propanoic acid(13),hupehcraboside(14),phlorizin(15)and 3-hydroxy phloridzin(16)by extensive spectroscopic analysis including 1D,2D NMR(HSQC,HMBC,1H-1H COSY,and ROESY),HRMS,ORD and CD spectra.The absolute configurations of 12,13 and 14 were assigned by comparing their calculated and experimental spectra,respectively.As a result,14 is a novel compound with unique skeleton,while 12,13 are two atropisomers and firstly isolated from natural resource.Meanwhile,an accurate and rapid method to simultaneously determine five main compounds,namely chlorogenic acid,protocatechuic acid,p-hydroxy-phenylpropionic acid,3-hydroxyl phloridzin andphlorizin in Hupeh crabapple,was firstly developed and validated.Subsequently,20 batches of smaples from different producing area were analized using this method.Results indicated that phlorizinwas the most abundant constituent with the content of 3.26～52.23 mg/g,while 3-hydroxyl phloridzin of 4.29～40.29 mg/g,p-hydroxy-phenylpropionic acid of 0.01～4.80 mg/g,chlorogenic of 0.021.75 mg/g and protocatechuic acid of 0.01～0.62 mg/g,respectively.Antioxidant activities of 12-15 were evaluated by measuring their DPPH free radical scavenging activities.Results indicated 12 and 13 shew significant antioxidant effects with the free radiacal clearance rates of 82.69%and 76.99%respectively. |
