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Differential Analysis of Organic Acids in Different Origins of Raspberry based on HPLC Combined with Multivariate Statistical Methods
LV Bolong, LI Guozhu
Differential Analysis of Organic Acids in Different Origins of Raspberry based on HPLC Combined with Multivariate Statistical Methods
To investigate the differences of eight organic acids in raspberry from different origins, a method for measuring the content of eight organic acids at the same time by HPLC was established, in order to provide a theoretical basis for some sour substances in raspberry. The raspberry herb of the same harvesting period was taken as the research object, the quantitative methods of eight organic acid components were established by HPLC, and the differences were analyzed and compared by cluster analysis (CA), principal component analysis (PCA) and orthogonal partial least squares-discriminant analysis (OPLS-DA). The results showed that the linear relationship of organic acids in the concentration range of 0.001-2.00 mg/mL was good, the limit of detection (LOD) were 0.326~1.114 μg/mL, the limit of quantification (LOQ) were 1.060~3.604 μg/mL, the average recovery rates were 97.40%-101.58%, and the relative standard deviation (RSD) were 1.23%-2.34%. This method determines the content of organic acids in raspberry of different origins, both cluster analysis and OPLS-DA can effectively distinguish some production areas according to provinces, and the effect of OPLS-DA is better than CA, the origins can be clustered into 7 categories according to the province: Zhejiang, Anhui, Jiangsu, Fujian, Jiangxi, Sichuan and Hunan. The PCA results showed that oxalic acid, tartaric acid, malic acid, shikimic acid, citric acid, succinic acid, and acetic acid were the main organic acids in raspberry herbs. HPLC was established to analyze the differences on the content of eight organic acids in raspberry at the first time, and the origins are wide, the results are reliable, which could be used for the quality evaluation and preliminary differentiation of organic acid components in raspberry of different origins.
raspberry / different origins / multivariate statistics / organic acids / HPLC {{custom_keyword}} /
表1 覆盆子药材产地及批号 |
编号 | 产地 | 批号 | 编号 | 产地 | 批号 | 编号 | 产地 | 批号 | ||
---|---|---|---|---|---|---|---|---|---|---|
1 | 江西婺源 | 20210522 | 11 | 安徽巢湖 | 20210520 | 21 | 福建南平 | 20210522 | ||
2 | 江西横峰 | 20210524 | 12 | 浙江千岛湖 | 20210518 | 22 | 福建大田 | 20210522 | ||
3 | 江西德兴 | 20210522 | 13 | 浙江衢州 | 20210523 | 23 | 福建政和 | 20210522 | ||
4 | 江西玉山 | 20210519 | 14 | 浙江三门 | 20210522 | 24 | 福建霞浦 | 20210522 | ||
5 | 安徽亳州 | 20210520 | 15 | 浙江兰溪 | 20210522 | 25 | 四川绵阳 | 20210520 | ||
6 | 安徽绩溪 | 20210428 | 16 | 浙江文成 | 20210522 | 26 | 四川广元 | 20210520 | ||
7 | 安徽歙县 | 20210525 | 17 | 浙江台州 | 20210522 | 27 | 江苏江阴 | 20210524 | ||
8 | 安徽广德 | 20210521 | 18 | 浙江宁波 | 20210522 | 28 | 江苏昆山 | 20210524 | ||
9 | 安徽泾县 | 20210521 | 19 | 浙江馨昌 | 20210522 | 29 | 广西柳州 | 20210522 | ||
10 | 安徽黄山 | 20210521 | 20 | 湖南衡阳 | 20210523 | 30 | 广西玉林 | 20210522 |
表2 8种有机酸标样的标准曲线方程、检测限和定量限 |
有机酸组分 | 保留时间/min | 标准曲线 | 相关系数r | 检测限/(μg/mL) | 定量限/(μg/mL) |
---|---|---|---|---|---|
草酸 | 3.212 | y=5000000x-4814.9 | 0.99995 | 0.645 | 2.080 |
酒石酸 | 3.546 | y=912187x+5323.2 | 0.99990 | 1.114 | 3.604 |
苹果酸 | 3.987 | y=795565x-5800.1 | 0.99975 | 0.326 | 1.060 |
莽草酸 | 4.254 | y=20000000x+72306 | 0.99999 | 0.579 | 1.730 |
柠檬酸 | 4.699 | y=790177x+8447.6 | 0.99990 | 0.401 | 1.209 |
乙酸 | 5.616 | y=479104x-2922.7 | 0.99999 | 0.682 | 2.060 |
琥珀酸 | 5.981 | y=464210x+4213.5 | 0.99985 | 0.558 | 1.601 |
富马酸 | 8.033 | y=9000000x+7577.4 | 0.99990 | 0.502 | 1.500 |
表3 8种有机酸的回收率和相对标准偏差 SRSD,n=6 |
有机酸组分 | 2.0 μg/mL | 100 μg/mL | 1000 μg/mL | |||||
---|---|---|---|---|---|---|---|---|
回收率% | SRSD/% | 回收率% | SRSD/% | 回收率% | SRSD/% | |||
草酸 | 101.2 | 1.24 | 99.01 | 2.11 | 98.61 | 1.68 | ||
酒石酸 | 98.29 | 1.58 | 97.59 | 1.24 | 99.90 | 1.33 | ||
苹果酸 | 98.45 | 1.73 | 99.64 | 1.49 | 98.41 | 1.28 | ||
莽草酸 | 98.90 | 1.47 | 100.81 | 1.45 | 101.58 | 1.23 | ||
柠檬酸 | 99.38 | 1.62 | 99.17 | 1.38 | 97.4 | 1.31 | ||
乙酸 | 100.42 | 1.49 | 98.74 | 1.32 | 101.02 | 1.43 | ||
琥珀酸 | 99.99 | 1.95 | 102.50 | 1.55 | 99.90 | 1.26 | ||
富马酸 | 99.83 | 2.22 | 98.26 | 1.46 | 100.26 | 2.34 |
表4 不同产地覆盆子中8种有机酸含量测定结果 mg/g,n=3 |
产地 | 缩写 | 草酸 | 酒石酸 | 苹果酸 | 莽草酸 | 乙酸 | 柠檬酸 | 琥珀酸 | 富马酸 |
---|---|---|---|---|---|---|---|---|---|
江西婺源 | JXWY | 1.089 | 2.319 | 1.669 | 0.052 | 0.702 | 3.226 | 2.16 | 0.004 |
江西横峰 | JXHF | 1.02 | 2.891 | 1.396 | 0.014 | 0.257 | 3.860 | 2.025 | 0.009 |
江西德兴 | JXDX | 1.054 | 2.605 | 1.533 | 0.033 | 0.480 | 3.543 | 2.092 | 0.006 |
江西玉山 | JXYS | 1.064 | 2.485 | 1.439 | 0.028 | 0.680 | 2.586 | 2.143 | 0.002 |
安徽亳州 | AHBZ | 1.405 | 1.731 | 2.177 | 0.098 | 0.565 | 3.122 | 2.714 | 0.002 |
安徽绩溪 | AHJX | 1.499 | 1.645 | 2.332 | 0.122 | 0.305 | 2.505 | 3.038 | 0.003 |
安徽歙县 | AHSX | 1.485 | 1.703 | 2.311 | 0.106 | 0.365 | 3.625 | 2.572 | 0.009 |
安徽广德 | AHGD | 1.491 | 1.684 | 2.316 | 0.11 | 0.211 | 4.588 | 2.323 | 0.003 |
安徽泾县 | AHJ | 1.544 | 1.546 | 2.395 | 0.13 | 0.266 | 2.843 | 2.633 | 0.004 |
安徽黄山 | AHHS | 1.525 | 1.584 | 2.371 | 0.126 | 0.415 | 3.011 | 2.849 | 0.003 |
安徽巢湖 | AHCH | 1.315 | 1.864 | 2.106 | 0.087 | 0.815 | 3.238 | 2.826 | 0.002 |
浙江千岛湖 | ZJQDH | 1.627 | 1.494 | 2.423 | 0.139 | 0.553 | 3.805 | 2.332 | 0.009 |
浙江衢州 | ZJQZ | 1.992 | 1.108 | 2.886 | 0.197 | 0.702 | 3.226 | 2.16 | 0.004 |
浙江三门 | ZJSM | 1.672 | 1.45 | 2.659 | 0.161 | 0.765 | 1.59 | 2.544 | 0.004 |
浙江兰溪 | ZJLX | 1.81 | 1.208 | 2.83 | 0.186 | 0.693 | 3.212 | 2.202 | 0.003 |
浙江文成 | ZJWC | 1.651 | 1.476 | 2.515 | 0.152 | 0.256 | 3.891 | 2.014 | 0.008 |
浙江台州 | ZJTZ | 1.702 | 1.353 | 2.763 | 0.175 | 0.567 | 3.032 | 2.482 | 0.009 |
浙江宁波 | ZJNB | 1.687 | 1.411 | 2.72 | 0.167 | 0.776 | 3.319 | 2.12 | 0.004 |
浙江馨昌 | ZJXC | 1.722 | 1.221 | 2.792 | 0.178 | 0.638 | 3.543 | 2.616 | 0.003 |
湖南衡阳 | HNHY | 1.448 | 1.724 | 2.206 | 0.104 | 0.932 | 1.712 | 1.582 | 0.010 |
福建南平 | FJNP | 1.078 | 2.457 | 1.449 | 0.032 | 0.716 | 3.539 | 2.585 | 0.002 |
福建大田 | FJDT | 1.094 | 2.252 | 1.744 | 0.062 | 0.428 | 3.229 | 2.792 | 0.008 |
福建政和 | FJZH | 1.084 | 2.366 | 1.617 | 0.037 | 0.402 | 2.886 | 2.631 | 0.005 |
福建霞浦 | FJXP | 1.219 | 2.118 | 1.987 | 0.075 | 0.569 | 2.569 | 2.609 | 0.006 |
四川绵阳 | SCMY | 1.144 | 2.239 | 1.868 | 0.07 | 0.921 | 2.347 | 1.728 | 0.006 |
四川广元 | SCGY | 1.255 | 2.03 | 2.033 | 0.08 | 0.661 | 3.031 | 1.66 | 0.006 |
江苏江阴 | JSJY | 1.349 | 1.78 | 2.12 | 0.091 | 0.445 | 1.856 | 1.692 | 0.002 |
江苏昆山 | JSKS | 1.361 | 1.743 | 2.142 | 0.095 | 0.731 | 3.831 | 2.366 | 0.006 |
广西柳州 | GXLZ | 1.228 | 2.056 | 2.016 | 0.078 | 0.532 | 1.757 | 3.001 | 0.004 |
广西玉林 | GXYL | 1.293 | 1.919 | 2.081 | 0.084 | 0.166 | 1.822 | 3.034 | 0.002 |
表5 不同产地覆盆子药材中有机酸的主成分分析 |
有机酸组分 | 主成分1 | 主成分2 | 主成分3 |
---|---|---|---|
草酸 | 0.987 | 0.030 | 0.063 |
酒石酸 | -0.988 | 0.038 | 0.029 |
苹果酸 | 0.994 | 0.002 | 0.006 |
莽草酸 | 0.995 | 0.031 | 0.012 |
乙酸 | 0.100 | 0.616 | -0.604 |
柠檬酸 | 0.043 | 0.049 | 0.801 |
琥珀酸 | 0.071 | -0.873 | -0.008 |
富马酸 | -0.044 | 0.594 | 0.542 |
特征根值 | 3.947 | 1.500 | 3.305 |
累计贡献率/% | 49.337 | 68.086 | 84.399 |
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