化学计量学二阶校正方法结合高效液相色谱用于蜂蜜中10种酚酸类物质的快速定量分析

doi:10.3724/SP.J.1123.2019.09013

摘要/Abstract

摘要：

利用化学计量学二阶校正方法结合高效液相色谱对枣花蜜中10种酚酸类物质的快速定量分析进行了研究。首先通过验证样本研究了所建立模型的准确性。结果显示：10种酚酸类物质的线性相关系数（R）为0.9982~0.9999，平均回收率为97.6%~101.1%，说明所建立的模型稳定可靠。其次，通过模拟蜂蜜试验，确定了固相萃取柱的种类及操作条件（HLB柱，酸化水淋洗，甲醇洗脱）。最后，利用模拟蜂蜜得到的最优条件结合化学计量学二阶校正方法，测定了枣花蜜中10种酚酸类物质的含量，并测得其加标回收率为62.1%~93.8%，考虑到目标分析物的种类较多，且蜂蜜基质极为复杂，该结果基本满足要求。另外，还利用统计与品质因子验证了试验方法的可靠性，结果令人满意。该方法具有简单、快速等优点，可用于复杂基质中多种目标分析物的同时定量分析。

关键词: 固相萃取, 高效液相色谱法, 酚酸类物质, 枣花蜜, 化学计量学

Abstract:

This study aims at the determination of ten phenolic acids in honey using high-performance liquid chromatography-diode array detection (HPLC-DAD) combined with a chemometric second-order calibration method. First, the accuracy of the model was investigated by analysis of the calibration samples and validation samples. Satisfactory results were obtained; the ten phenolic acids exhibited good linearity, and the correlation coefficient (R) was in the range 0.9982-0.9999, with average recoveries of 97.6% to 101.1%, which proved that the model was stable and reliable. Second, the types and operating conditions of the SPE columns were determined by experiments using simulated honey; HLB column was selected, washed with 0.1%(v/v) formic acid aqueous solution, and eluted with methanol. Finally, the phenolic acid contents in jujube flower honey were determined under the optimum conditions for simulated honey. The recoveries of the analytes were 62.1%-93.8%, which met the requirements considering the variety of target analytes and the complexity of the honey substrate. In addition, the efficiency of the method was validated by figures of merit and statistical parameters, including sensitivity (SEN), selectivity (SEL), root-mean-square error of prediction (RMSEP), limit of detection (LOD), and limit of quantification (LOQ); the obtained results were satisfactory. This method is simple, fast, and accurate, and it can be used for the quantitative analysis of the target analytes in complex substrates.

Key words: solid phase extraction (SPE), high-performance liquid chromatography (HPLC), phenolic acids, jujube flower honey, chemometrics

张晓华, 吕敏明, 郑晶晶, 母淑婷, 刘攀华, 陈征. 化学计量学二阶校正方法结合高效液相色谱用于蜂蜜中10种酚酸类物质的快速定量分析[J]. 色谱, 2020, 38(5): 572-580.

ZHANG Xiaohua, LÜ Minming, ZHENG Jingjing, MU Shuting, LIU Panhua, CHEN Zheng. Fast high-performance liquid chromatography quantification of ten phenolic acids in honey using chemometric second-order calibration method[J]. Chinese Journal of Chromatography, 2020, 38(5): 572-580.

图/表 8

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No.	GA	3, 4-DA	CHA	p-HA	CA	SA	p-CA	FA	AA	CIA
GA: gallic acid; 3, 4-DA: 3, 4-dihydroxybenzoic acid; CHA: chlorogenic acid; p-HA: p-hydroxybenzoic acid; CA: caffeic acid; SA: syringic acid; p-CA: p-coumaric acid; FA: ferulic acid; AA: abscisic acid; CIA: cinnamic acid. C01-C06: calibration sample 1-calibration sample 6. V01-V05: validation sample 1-validation sample 5.
C01	0.00	0.00	4.38	0.00	4.40	0.00	0.00	5.12	0.00	5.48
C02	2.76	0.44	3.94	0.44	3.96	4.32	0.43	4.61	0.44	4.93
C03	11.04	1.74	2.63	1.76	2.64	0.86	1.72	3.07	1.76	3.29
C04	16.56	2.62	1.75	2.64	1.76	1.30	2.58	2.05	2.64	2.19
C05	22.08	3.49	0.88	3.52	0.88	1.73	3.44	1.02	3.52	1.10
C06	27.60	4.36	0.00	4.40	0.00	2.16	4.30	0.00	4.40	0.00
V01	5.52	0.87	3.50	0.88	3.52	0.43	0.86	4.10	0.88	4.38
V02	8.28	1.31	3.07	1.32	3.08	3.89	1.29	3.58	1.32	3.84
V03	13.80	2.18	2.19	2.20	2.20	3.46	2.15	2.56	2.20	2.74
V04	19.32	3.05	1.31	3.08	1.32	3.02	3.01	1.54	3.08	1.64
V05	24.84	3.92	0.44	3.96	0.44	2.59	3.87	0.51	3.96	0.55

No.	GA	3, 4-DA	CHA	p-HA	CA	SA	p-CA	FA	AA	CIA
GA: gallic acid; 3, 4-DA: 3, 4-dihydroxybenzoic acid; CHA: chlorogenic acid; p-HA: p-hydroxybenzoic acid; CA: caffeic acid; SA: syringic acid; p-CA: p-coumaric acid; FA: ferulic acid; AA: abscisic acid; CIA: cinnamic acid. C01-C06: calibration sample 1-calibration sample 6. V01-V05: validation sample 1-validation sample 5.
C01	0.00	0.00	4.38	0.00	4.40	0.00	0.00	5.12	0.00	5.48
C02	2.76	0.44	3.94	0.44	3.96	4.32	0.43	4.61	0.44	4.93
C03	11.04	1.74	2.63	1.76	2.64	0.86	1.72	3.07	1.76	3.29
C04	16.56	2.62	1.75	2.64	1.76	1.30	2.58	2.05	2.64	2.19
C05	22.08	3.49	0.88	3.52	0.88	1.73	3.44	1.02	3.52	1.10
C06	27.60	4.36	0.00	4.40	0.00	2.16	4.30	0.00	4.40	0.00
V01	5.52	0.87	3.50	0.88	3.52	0.43	0.86	4.10	0.88	4.38
V02	8.28	1.31	3.07	1.32	3.08	3.89	1.29	3.58	1.32	3.84
V03	13.80	2.18	2.19	2.20	2.20	3.46	2.15	2.56	2.20	2.74
V04	19.32	3.05	1.31	3.08	1.32	3.02	3.01	1.54	3.08	1.64
V05	24.84	3.92	0.44	3.96	0.44	2.59	3.87	0.51	3.96	0.55

No.	Analyte	Linear equation	R	Average recovery/%
y: the relative concentration values of each analyte in calibration samples; x: the nominal concentrations in calibration samples.
1	GA	y=8.43x+2.40	0.9999	99.0
2	3, 4-DA	y=10.99x+3.20	0.9993	100.6
3	CHA	y=5.77x+0.38	0.9993	99.4
4	p-HA	y=11.54x+2.30	0.9992	101.0
5	CA	y=3.93x+1.18	0.9990	98.4
6	p-CA	y=1.55x+2.15	0.9998	97.6
7	SA	y=16.68x+0.68	0.9982	101.1
8	FA	y=5.02x+0.91	0.9987	98.6
9	AA	y=8.12x+0.99	0.9998	99.4
10	CIA	y=0.14x+0.63	0.9998	99.0

No.	Analyte	Linear equation	R	Average recovery/%
y: the relative concentration values of each analyte in calibration samples; x: the nominal concentrations in calibration samples.
1	GA	y=8.43x+2.40	0.9999	99.0
2	3, 4-DA	y=10.99x+3.20	0.9993	100.6
3	CHA	y=5.77x+0.38	0.9993	99.4
4	p-HA	y=11.54x+2.30	0.9992	101.0
5	CA	y=3.93x+1.18	0.9990	98.4
6	p-CA	y=1.55x+2.15	0.9998	97.6
7	SA	y=16.68x+0.68	0.9982	101.1
8	FA	y=5.02x+0.91	0.9987	98.6
9	AA	y=8.12x+0.99	0.9998	99.4
10	CIA	y=0.14x+0.63	0.9998	99.0

Analyte	Added/ (μg/g)	HLB		PSD		MAX
Analyte	Added/ (μg/g)	Found/(μg/g)	Recovery/%	Found/(μg/g)	Recovery/%	Found/(μg/g)	Recovery/%
GA	3.45	3.53	102.2	1.92	55.7	3.52	102.1
3, 4-DA	0.55	0.46	82.8	0.37	67.3	0.43	78.2
CHA	0.55	0.50	90.6	0.47	85.5	0.01	1.8
p-HA	0.55	0.50	90.9	0.46	83.6	0.46	83.6
CA	0.86	0.61	70.9	0.61	70.9	0.62	72.1
SA	0.54	0.44	81.5	0.37	68.5	0.36	66.7
p-CA	0.84	0.54	64.3	0.53	63.1	0.52	61.9
FA	1.00	0.72	72.0	0.70	70.0	0.72	72.0
AA	0.55	0.45	82.6	0.43	78.2	0.43	78.2
CIA	0.69	0.53	76.8	0.52	75.4	0.53	76.8