6 7 Vol. 6 No. 7 2015 7 Journal of Food Safety and Quality Jul., 2015 Ⅰ Ⅱ 钱 勇 1*, 许纪锋 1, 诸晨 1, 沈君子 1, 金金花 2, 梅利中 2, 谢天培 3 (1., 201203; 2., 201308; 3., 201203) 摘要 : 目的 (high performance liquid chromatography, HPLC), 方法 90%, Ultimate XB-C 18 (4.6 mm 250 mm, 5 μm), : =32:68 (V:V)(0.1% ), 1.0 ml/min, 35, 440 nm 结果 15~300 μg/ml (r 2 =0.9996), 96.6%, RSD 1.69%; 12~240 μg/ml (r 2 =0.9998), 96.1%, RSD 1.58%, 结论, 关键词 : ; Ⅰ; Ⅱ; ; Determination of crocusⅠand crocusⅡin saffron QIAN Yong 1*, XU Ji-feng 1, ZHU Chen 1, SHEN Jun-Zi 1, JIN Jin-Hua 2, MEI Li-Zhong 2, SXIE Tian-Pei 3 (1. Shanghai Standard Biotech Co., Ltd., Shanghai 201203, China; 2. Shanghai Huapu Medicine Decoction Pieces Co., Ltd., Shanghai 201308, China; 3. Shanghai 100 Standard Testing Co., Ltd., Shanghai 201203, China) ABSTRACT: Objective To develop a method for determination of crocus Ⅰ and crocus Ⅱ in saffron by high performance liquid chromatography (HPLC), and compare the content of crocusⅠand crocus Ⅱ in different saffron. Methods The saffron was extracted by ultrasonic in 90% ethanol solution. Separations of extraction solution were carried out on an Ultimate XB-C 18 (4.6 mm 250 mm, 5 μm). The mobile phase(volume rate)was ACN:H 2 O = 32:68 (V:V) (0.1% methanoic acid), the flow was 1.0 ml/min, column temperature was 35, and the detection wavelength was 440 nm. Results Crocus Ⅰ had a good linearity with the peak areas in 15~300 μg/ml (r 2 =0.9996). The average recovery was 96.6% and the RSD was 1.69%. Crocus Ⅱ had a good linearity with the peak areas in 12~240 μg/ml(r 2 =0.9998). The average recovery was 96.1% and the RSD was 1.58%. The difference of content of crocus Ⅰ and crocus Ⅱ was obvious in different cultivated fields. Conclusion This method is simple, sensitive, specific, and reproducible, which can be used as the method for measuring the content of crocus Ⅰ and crocus Ⅱ in saffron. KEY WORDS: saffron; crocus Ⅰ; crocus Ⅱ; high performance liquid chromatography; determination of content 基金项目 : (14401903200) Fund: Supported by the Science and Technology Plan of STCSM (14401903200) * 通讯作者 :,, E-mail: qianyong2232@163.com *Corresponding author: QIAN Yong, Engineer, Shanghai Standard Biotech Co., Ltd., Building 2, Room 103, No.720, Cailun Road, Shanghai 201203, China. E-mail: qianyong2232@163.com
7, : Ⅰ Ⅱ 2823 1 引言 (Crocus sativus L.),, B1, ; [1-3], α- [4-7], 1,,,,, [8-10], (high performance liquid chromatography, HPLC) [11-16],, HPLC, [11,12,16] 1 Ⅰ(1) Ⅱ(2) Fig. 1 Structure of crocusⅠ(1) and crocus Ⅱ (2)
2824 6, Ⅱ, Ⅰ Ⅱ,, 2 材料与方法 2.1 材料与试剂 ( 98.5%, ), (, ),, (, ), (, ) 2.2 仪器与设备 Agilent1260, Chemstation ( ); ( ); ELGA ( ); HS-800D ( ); Fresco-21 ( ) 2.3 方法过程 2.3.1 色谱条件 : Ultimate XB- C18(4.6 mm 250 mm, 5 μm), : : =32:68 (V:V)( 0.1% ) 1.0 ml/min, 440 nm, 35 2.3.2 对照品溶液的制备 15.0 mg 12.0 mg 25 ml,,, 0.60 mg/ml, 0.48 mg/ml 2 Fig. 2 Liquid chromatogram of saffron (1: Ⅰ, 2: Ⅱ; A, B ) (1: crocusⅠ, 2: crocusⅡ; A: reference standards of crocusⅠand crocusⅡ; B: extract sample of saffron)
7, : Ⅰ Ⅱ 2825 2.3.3 供试药材溶液的制备 0.1 g,, 100 ml, 90% 60 ml, 40 20 min, 2,,,, 2 ml 10 ml,,, 0.45 μm, 3 结果与分析 3.1 线性范围的考察, 15 30 60 150 300 μg/ml 5, 10 μl,, 12 24 48 120 240 μg/ml 5, 10 μl, (Y), (X), : Y=36655X+8.3597(r 2 =0.9996), 15~300 μg/ml ; : Y=12606X-0.0905, (r 2 =0.9998) 12~240 μg/ml 3.2 精密度试验, 6, 10 μl, RSD 0.65%, RSD 0.95%, 3.3 稳定性试验, 0 1 2 4 8 12 24 h, 7, 10 μl, RSD 0.55%, RSD 0.75%, 24 h 3.4 重现性试验 5,,, 10 μl,, 79.5 mg/g 28.6 mg/g, RSD 1.75% 1.42% 3.5 加样回收率试验 6,, 0.5 1 2 ml 2,,, 10μL, 96.6% 96.1%, RSD 1.69% 1.58% 3.6 不同产地的西红花苷 Ⅰ 和西红花苷 Ⅱ 含量测定,,, 10 μl, 2,, 1 表 1 Table 1 不同产地西红花中西红花苷 Ⅰ 和西红花苷 Ⅱ 的含量 (n=3) Content of crocusⅠand crocusⅡ in different saffron (n=3) Ⅰ(%) Ⅱ(%) Ⅰ Ⅱ (%) RSD(%)(Ⅰ Ⅱ ) XHH-1 7.97 2.87 10.84 1.57 XHH-2 5.65 1.64 7.29 1.78 XHH-3 2.14 0.73 2.87 1.15 XHH-4 0.85 0.37 1.22 0.98 XHH-5 8.35 3.10 11.45 1.59 XHH-6 6.20 2.65 8.85 1.46
2826 6 Ⅰ(%) Ⅱ(%) Ⅰ Ⅱ (%) RSD(%)(Ⅰ Ⅱ ) 续表 1 XHH-7 14.10 4.70 18.80 1.36 XHH-8 3.28 1.15 4.43 0.95 XHH-9 10.20 3.65 13.85 1.85 XHH-10 12.51 4.24 16.75 1.25 XHH-16 8.56 3.10 11.66 0.87 XHH-11 14.65 4.74 19.39 1.46 XHH-12 11.34 4.87 16.21 1.75 XHH-13 23.47 8.92 32.39 1.55 XHH-14 10.25 3.74 13.99 1.11 XHH-15 7.54 2.64 10.18 0.83 XHH-16 13.26 4.85 18.11 0.98 XHH-18 6.32 1.75 8.07 1.67 XHH-19 4.34 1.58 5.92 1.32 XHH-20 7.32 2.54 9.86 1.72 XHH-21 9.32 3.46 12.78 1.82 4 结论与讨论 4, 21,,, [17],,, 10.0%,,,, ( 2~4 ),, [11,12,16],,,, 参考文献 [1]. ( )[M]. :, 2010. Chinese Pharmacopoeia Commission. Pharmacopoeia of the People's Republic of China (First Volume) [M]. Beijing: Chinese Medical Science and Technology Press, 2010. [2] Amin A, Hamza AA, Bajbouj K, et al. Saffron: a potential candidate for a novel anticancer drug against hepatocellular carcinoma [J]. Hepatology, 2011, 54(3): 857 867. [3] Tavakkol, Afshari J, Brook A, et al. Study of cytotoxic and apoptogenie properties of saffron extract in human cancer cell lines [J]. Food Chem Toxical, 2008, 46(11): 3443 3447. [4] Bajbouj K, Schulze-Luehrmann J, Diermeier S, et a1. The anti-cancer effect of saffron in two p53 isogenic colorectal cancer cell lines [J]. BMC Compl Altem Med, 2012, 12: 69. [5] Aung HH, Wang CZ, Ni M, et a1. Crocin from crocus sativus
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