薄層熒光掃描法測定花生根中白藜蘆醇的含量

韓小麗,張士真,呼亞旭,李明靜＊

(1.河南省醫(yī)藥學(xué)校,河南開封475001; 2.商丘市技師學(xué)院,河南商丘476000;3.河南大學(xué)化學(xué)化工學(xué)院,河南開封475004)

薄層熒光掃描法測定花生根中白藜蘆醇的含量

韓小麗1,張士真2,呼亞旭3,李明靜3＊

(1.河南省醫(yī)藥學(xué)校,河南開封475001; 2.商丘市技師學(xué)院,河南商丘476000;3.河南大學(xué)化學(xué)化工學(xué)院,河南開封475004)

在硅膠 G鋁箔板上,以V(氯仿)∶V(丙酮)∶V(甲酸)=8∶1∶0.2的溶液為展開劑,狹縫尺寸為3 mm×0.45 mm,檢測波長為335 nm,采用薄層熒光掃描法測定花生根中白藜蘆醇的含量.結(jié)果表明:白藜蘆醇在21.6～108.0 ng范圍內(nèi)線性關(guān)系良好,回歸方程為 Y=-664.701+12.666X,R=0.996 9,花生根中白藜蘆醇的含量為358.89μg/g.

薄層熒光掃描法;白藜蘆醇;花生根

白藜蘆醇(resveratrol)是一種具有生物活性的多酚類化合物,其化學(xué)名稱為3,4′,5-三羥基二苯乙烯.目前至少已經(jīng)在21科31屬72種植物中發(fā)現(xiàn)了白藜蘆醇的存在,如豆科的落花生屬、葡萄科的葡萄屬,決明屬、百合科的藜蘆屬、槐屬[1]等.由于它具有抗氧化,抑制癌細(xì)胞增殖,抑制血小板活性,消炎等多種藥理活性[2-4],因此被廣泛應(yīng)用于醫(yī)藥、食品、化妝品等行業(yè).

目前,開發(fā)利用富含白藜蘆醇的保健植物愈來愈受到人們的關(guān)注,有關(guān)從葡萄皮、虎杖中提取白藜蘆醇的研究已有大量的文獻(xiàn)報道[5-7],而花生根中白藜蘆醇的研究報道較少,而且主要集中于用 HPLC法測定白藜蘆醇的含量[8-10].花生根為花生的不可食部分,如能充分利用,既可以提高花生種植的附加值,又可避免環(huán)境污染,對農(nóng)產(chǎn)品資源開發(fā)與利用將有重要的意義.作者用薄層熒光掃描法對花生根中白藜蘆醇的含量進(jìn)行了測定,為綜合利用提供參考.

1 實(shí)驗(yàn)部分

1.1 儀器與材料

970CRT熒光分光光度計(jì)(上海精密科學(xué)儀器有限公司),CAMAG TLC SANNER 3型薄層掃描儀(瑞士),CAMAG LINOMAT5半自動點(diǎn)樣儀(瑞士),鋁箔板(德國,20 mm×20 mm×5 mm),白藜蘆醇對照品(購自中科院昆明植物所植化室),其他試劑均為分析純.將花生根(采自河南開封近郊)洗凈、晾干、粉碎后,備用.

1.2 色譜及掃描條件

展開劑:V(氯仿)∶V(丙酮)∶V(甲酸)=8∶1∶0.2;上行展開,展距5 cm;單光束反射式熒光線性掃描,激發(fā)波長335 nm;狹縫尺寸3 mm×0.45 mm,濾光片 K400;掃描速度20 mm·s-1.

2 結(jié)果與討論

2.1 對照品溶液的制備

精密稱取白藜蘆醇對照品適量,用乙酸乙酯溶解,定容至25 mL容量瓶中,配制成濃度為0.108 0 g/L的對照品溶液備用.

2.2 樣品溶液的制備

取花生根粗粉3 g,精密稱定,置于索氏提取器中,用90 mL 95%的甲醇冷浸,分三次加入,冷浸24 h后索氏提取3 h,水浴溫度75℃.提取液旋轉(zhuǎn)蒸至醇干.加入等體積石油醚萃取三次,萃余液用等體積乙酸乙酯萃取三次,收集有機(jī)相,旋轉(zhuǎn)蒸干.用乙酸乙酯溶解,轉(zhuǎn)移至10 mL容量瓶中定容,待測.

2.3 標(biāo)準(zhǔn)曲線的考察

用半自動點(diǎn)樣儀對白藜蘆醇對照品溶液進(jìn)行點(diǎn)樣,點(diǎn)樣量分別為0.1,0.2,0.3,0.4,0.5,0.6,0.7,0.8,0.9,1.0μL,點(diǎn)于同一塊硅膠 G鋁箔板上進(jìn)行檢測,以白藜蘆醇峰面積(Y)為縱坐標(biāo),進(jìn)樣體積(X)為橫坐標(biāo),繪制標(biāo)準(zhǔn)曲線,回歸方程為:Y=-664.701+12.666X,R=0.996 9;在21.6 ng～108.0 ng范圍內(nèi)線性關(guān)系良好.

2.4 樣品含量測定

精密稱取等量的花生根粗粉(3份),按照2.2項(xiàng)制備樣品溶液,在1.2項(xiàng)中的色譜和掃描條件下進(jìn)行測定,測得樣品含量平均值為358.89μg/g,RSD為1.49%.測定結(jié)果見表1.

2.5 精密度試驗(yàn)

精密吸取5份0.3μL的對照品溶液,分別點(diǎn)于同一塊硅膠 G鋁箔板上,展開,掃描,測得峰面積值分別為3 622.57,3 682.60,3 529.69,3 524.13,3 816.76.平均值為3 635.15,RSD為3.34%.

2.6 重現(xiàn)性試驗(yàn)

取2.4項(xiàng)中樣品2,分別吸取0.3μL在同一塊硅膠 G鋁箔板上點(diǎn) 5個點(diǎn),掃描,測得峰面積值分別為4 958.45,

4 817.56,4 818.45,5 120.46,4 996.81,平均值為4 942.35,RSD為2.60%.

2.7 穩(wěn)定性試驗(yàn)

取上述樣品2,放置5 h、10 h、15 h和24 h后分別進(jìn)行測定,測得樣品中白藜蘆醇含量分別為363.98,363.69,359.77,359.44,359.34μg/g,平均值為361.24μg/g,RSD為0.66%.可知樣品在冰箱內(nèi)(6℃)存放24 h后白藜蘆醇含量基本穩(wěn)定.

2.8 回收率試驗(yàn)

取2.4項(xiàng)中樣品溶液5.0 mL,加入0.108 0 g/L的對照品溶液0.5 mL,用乙酸乙酯溶解,并定容至10 mL容量瓶中,在1.2項(xiàng)中的色譜和掃描條件下測定,回收率分別為103.86%,105.08%,105.52%,平均回收率為104.82%,RSD=0.80%(n=3).

2.9 熒光激發(fā)波長的選擇

白藜蘆醇在366 nm的紫外光照射下能產(chǎn)生熒光,但這并不是最佳的熒光激發(fā)波長.通過熒光分光光度計(jì)的檢測得知,最佳激發(fā)波長為335 nm,熒光發(fā)射波長為375 nm,本試驗(yàn)中的熒光為Stokes熒光.

71.

表1 樣品含量測定結(jié)果(n=3)Table 1 Results of sample determination

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Determination of Resveratrol in Peanut Root by Thin Layer Chromatography Fluorescence Scanning

HAN Xiao-li,ZHANG Shi-zhen,HU Ya-xu,LI Ming-jing
(1.Henan Tharmaceutital School,Kaif eng475001,Henan,China; 2.College of Shangqiu Technician,Shangqiu476000,Henan,China; 3.College of Chemistry and Chemical Engineering,Henan University,Kaif eng475004,Henan,China)

The content of resveratrol was determined by thin layer chromatography fluorescence scanning on silica-gel G aluminium foil plate.The developing solvent was composed by chloroform,acetone and formic acid(8∶1∶0.2).The slit size was 3 mm×0.45 mm and the detection wavelength was 335 nm.The results showed that the calibration curves were linear in the range of 21.6～108.0 ng(R=0.996 9),and the content of resveratrol in peanut root was 358.89 μg/g.

thin layer chromatography fluorescence scanning;resveratrol;peanut root

O 652.63

A

1008-1011(2010)05-0088-02

2010-05-20.

韓小麗(1982-)女,碩士生,研究方向:天然產(chǎn)物及中草藥化學(xué).＊

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