竹叶提取物中7种黄酮成分含量测定及抗氧化分析

doi:10.3724/SP.J.1123.2024.01008

摘要/Abstract

摘要：

为研究不同竹叶提取物中黄酮成分含量及其与抗氧化活性之间的关系,以2种市售竹叶提取物以及以毛竹等7种竹种的竹叶为材料制备的竹叶提取物共9个样品为研究对象,建立了同时测定竹叶提取物中7种黄酮成分(荭草苷、异荭草苷、牡荆苷、异牡荆苷、苜蓿素、木犀草素和木犀草苷)的高效液相色谱(HPLC)方法,采用SymmetryShield^TM RP8色谱柱(250 mm×4.6 mm, 5 μm),以乙腈为流动相A, 0.5%(v/v)乙酸水溶液为流动相B进行梯度洗脱。采用1,1-二苯基-2-三硝基苯肼(DPPH)和羟基自由基清除试验评价竹叶提取物的抗氧化活性,以半抑制浓度(IC₅₀)为评价指标,以抗氧化剂2,6-二叔丁基对甲酚(BHT)和叔丁基对苯二酚(TBHQ)为阳性对照,并利用Person相关性分析各黄酮成分含量与抗氧化的关系。结果表明:建立的HPLC测定方法准确可靠,适用于竹叶提取物中黄酮含量的测定。竹叶提取物中7种黄酮含量在14.97~183.94 mg/g范围内,毛竹中7种黄酮总含量最高,为183.94 mg/g。不同竹种间7种黄酮成分的含量存在显著差异,荭草苷、异荭草苷和牡荆苷含量均在毛竹中最高,分别为38.45、101.30和9.42 mg/g。竹叶提取物清除DPPH自由基的IC₅₀值在78.23~179.41 mg/L范围内,清除羟基自由基的IC₅₀值在203.48~1250.81 mg/L范围内。毛竹叶提取物清除DPPH和羟基自由基的IC₅₀值分别为78.23 mg/L和203.48 mg/L,其抗氧化活性最强,具有开发应用价值。相关性分析结果表明,荭草苷、异荭草苷含量与竹叶提取物抗氧化活性密切相关。

关键词: 高效液相色谱, 竹叶提取物, 黄酮, 抗氧化

Abstract:

The flavonoid contents of different bamboo-leaf extracts and their relationships to antioxidant activity were investigated in this study by preparing nine samples using two commercially available bamboo-leaf extract products and seven bamboo-leaf extracts such as Phyllostachys edulis. A high performance liquid chromatography (HPLC) method was established to determine seven flavonoid components (orientin, isoorientin, vitexin, isovitexin, tricin, luteolin and luteoloside) in these samples, which were separated using a SymmetryShield^TM RP8 column (250 mm×4.6 mm, 5 μm) under gradient-elution conditions using acetonitrile as mobile phase A and 0.5% (v/v) acetic acid aqueous solution as mobile phase B. The antioxidant activities of the samples were evaluated using the 1,1-diphenyl-2-picrylhydrazyl (DPPH) and hydroxyl radical-scavenging assays, with half inhibitory concentration (IC₅₀) as an indicator and the butylated hydroxytoluene (BHT) and tert-butylhydroquinone (TBHQ) antioxidants as positive controls. Pearson correlation was then used to analyze the relationship between flavonoid content and antioxidant activity. The HPLC method was found to be accurate and reliable for determining the flavonoid contents of the bamboo-leaf extracts. The seven flavonoids were well separated, and good linear relationships were exhibited (correlation coefficients (R²)≥0.9990). Furthermore, the contents of the seven flavonoids in the bamboo-leaf extracts ranged from 14.97 to 183.94 mg/g, with the highest content of 183.94 mg/g recorded for Phyllostachys edulis. The bamboo species exhibited significantly different flavonoid contents, with Phyllostachys edulis showing the highest orientin, isoorientin, and vitexin levels of 38.45, 101.30, and 9.42 mg/g, respectively. Moreover, the bamboo-leaf extracts exhibited IC₅₀ values of 78.23-179.41 mg/L for DPPH-radical-scavenging, while values of 203.48-1250.81 mg/L were recorded for hydroxyl radicals. The Phyllostachys edulis leaf extract exhibited the strongest antioxidant activity, with the lowest IC₅₀ values of 78.23 and 203.48 mg/L for DPPH and hydroxyl, respectively; it showed greatly significant for the further development and application of Phyllostachys edulis. Finally, the relationships between flavonoid content and the DPPH- and hydroxyl-radical-scavenging activities (based on the IC₅₀ values) were correlated, which revealed that the orientin and isoorientin contents are closely related to the antioxidant activities of the bamboo-leaf extracts. Consequently, the orientin and isoorientin contents can be used as indicators for evaluating the antioxidant activities of bamboo-leaf extracts.

Key words: high performance liquid chromatography (HPLC), bamboo-leaf extracts, flavonoid, antioxidant

中图分类号:

O658

顾丽玲, 姚曦, 安容苗, 郭雪峰. 竹叶提取物中7种黄酮成分含量测定及抗氧化分析[J]. 色谱, 2024, 42(10): 972-978.

GU Liling, YAO Xi, AN Rongmiao, GUO Xuefeng. Determination and antioxidant analysis of seven flavonoids in bamboo-leaf extracts[J]. Chinese Journal of Chromatography, 2024, 42(10): 972-978.

图/表 6

参考文献

[1]	Li Z Y, Trinh T L, Li N, et al. World Bamboo and Rattan, 2020, 18(4): 1
[1]	李智勇, Trinh T L, 李楠, 等. 世界竹藤通讯, 2020, 18(4): 1
[2]	Yang S, Wu W G, Guo L, et al. Journal of Chinese Medicinal Materials, 2022, 45(3): 743
[2]	杨帅, 吴卫刚, 郭亮, 等. 中药材, 2022, 45(3): 743
[3]	Yue Y D, Cao H Q, Tang F. Journal of Anhui Agricultural University, 2007, 105(3): 328
[3]	岳永德, 操海群, 汤锋. 安徽农业大学学报, 2007, 105(3): 328
[4]	Zhang Y L, Zhao J C, Yang Z Y, et al. Science and Technology of Food Industry, 2022, 43(9): 93
[4]	张亚兰, 赵建诚, 杨振亚, 等. 食品工业科技, 2022, 43(9): 93
[5]	Yu Y, Li Z M, Cao G T, et al. J Food Sci, 2019, 84(6): 1609
[6]	Zhang S, Chen J, Sun A D, et al. Food Agric Immunol, 2014, 25(3): 386
[7]	Nur A E, Umar S, Supriyadi S. Key Engineering Materials, 2021, 884: 256
[8]	Chen Y L, Chen C, Lai K H, et al. J Funct Foods, 2023, 101: 105407
[9]	Yang C, Lv Y F, Liang D, et al. Drug Res (Stuttg), 2015, 65(12): 668
[10]	Li Y. [MS Dissertation]. Nanchang: Jiangxi Agricultural University, 2022
[10]	李莹.[硕士学位论文]. 南昌: 江西农业大学, 2022
[11]	Gu Y P, Fan X, Xiao H R, et al. Nutrients, 2022, 14(4): 793
[12]	Chen D D, Guo X F, Zhao L, et al. Chemistry and Industry of Forest Products, 2019, 39(5): 108
[12]	陈丹丹, 郭雪峰, 赵蕾, 等. 林产化学与工业, 2019, 39(5): 108
[13]	Shao S Y. [MS Dissertation]. Beijing: Chinese Academy of Forestry, 2020
[13]	邵思越.[硕士学位论文]. 北京: 中国林业科学研究院, 2020
[14]	Tang F, Wu J Y, Zhang J, et al. Journal of Jilin University (Science Edition), 2013, 51(4): 724
[14]	汤峰, 吴静义, 张剑, 等. 吉林大学学报(理学版), 2013, 51(4): 724
[15]	Guo J, Wang H R, Shen Z Y, et al. Chinese Traditional Patent Medicine, 2019, 41(11): 2688
[15]	郭静, 王浩然, 沈周媛, 等. 中成药, 2019, 41(11): 2688
[16]	Li X B, Tao W Q, Xun H, et al. Rev Bras Farmacogn, 2021, 31: 347
[17]	Ma N H, Guo J, Xu C S H, et al. Molecules, 2020, 25(2): 409
[18]	Cui J, Yue Y D, Tang F, et al. JPC-J Planar Chromat, 2011, 24(5): 394
[19]	Wang P, Jiang D, Mei J, et al. China Food Additives, 2020, 31(12): 70
[19]	王鹏, 江丹, 梅菊, 等. 中国食品添加剂, 2020, 31(12): 70
[20]	Yuan F J, Yu L, Zhang M, et al. Guangxi Sciences, 2022, 29(6): 1141
[20]	袁凤娟, 于璐, 张萌, 等. 广西科学, 2022, 29(6): 1141
[21]	Zhang A L, Shen Y W, Cen M P, et al. Ind Crops Prod, 2019, 133: 111

Component	Linear equation	Linear range/(mg/L)	R²	LOD/(mg/L)	LOQ/(mg/L)
Orientin	y=15836x-51328	1.13-170	0.9993	0.40	0.90
Isoorientin	y=26962x-115503	1.20-180	0.9990	0.48	1.20
Vitexin	y=21948x+8583.1	1.53-230	0.9996	0.40	1.10
Isovitexin	y=30338x-23490	1.13-170	0.9999	0.25	0.80
Luteoloside	y=15355x+18309	1.00-110	0.9992	0.40	0.80
Tricin	y=24059x+19988	0.73-110	0.9993	0.21	0.70
Luteolin	y=26111x-70153	1.20-120	0.9990	0.20	0.50

Component	Linear equation	Linear range/(mg/L)	R²	LOD/(mg/L)	LOQ/(mg/L)
Orientin	y=15836x-51328	1.13-170	0.9993	0.40	0.90
Isoorientin	y=26962x-115503	1.20-180	0.9990	0.48	1.20
Vitexin	y=21948x+8583.1	1.53-230	0.9996	0.40	1.10
Isovitexin	y=30338x-23490	1.13-170	0.9999	0.25	0.80
Luteoloside	y=15355x+18309	1.00-110	0.9992	0.40	0.80
Tricin	y=24059x+19988	0.73-110	0.9993	0.21	0.70
Luteolin	y=26111x-70153	1.20-120	0.9990	0.20	0.50

Component	Original/ (mg/g)	Added/ (mg/g)	Measured/ (mg/g)	Recovery/ %	RSD/ %
Orientin	38.55	17.00	54.36	92.96	4.06
		26.98	64.70	96.74	1.66
		53.97	91.20	97.57	1.28
Isoorientin	94.80	18.00	112.10	95.90	3.91
		28.57	122.93	98.43	1.04
		57.14	151.42	99.08	0.49
Vitexin	8.94	23.00	31.38	97.57	1.56
		36.51	45.42	99.94	0.50
		73.02	82.08	100.18	0.81
Isovitexin	30.11	17.00	46.97	99.18	2.55
		26.98	55.43	93.84	2.35
		53.97	82.50	97.09	0.41
Luteoloside	2.92	11.00	13.86	99.42	2.05
		17.46	20.43	100.29	1.60
		34.92	36.98	97.53	0.92
Tricin	0.11	11.00	11.10	99.92	1.21
		17.46	18.66	106.25	1.00
		34.92	35.51	101.37	0.86
Luteolin	0.17	18.00	19.68	108.35	1.55
		28.57	30.13	104.84	1.40
		57.14	62.65	109.33	1.24

Component	Original/ (mg/g)	Added/ (mg/g)	Measured/ (mg/g)	Recovery/ %	RSD/ %
Orientin	38.55	17.00	54.36	92.96	4.06
		26.98	64.70	96.74	1.66
		53.97	91.20	97.57	1.28
Isoorientin	94.80	18.00	112.10	95.90	3.91
		28.57	122.93	98.43	1.04
		57.14	151.42	99.08	0.49
Vitexin	8.94	23.00	31.38	97.57	1.56
		36.51	45.42	99.94	0.50
		73.02	82.08	100.18	0.81
Isovitexin	30.11	17.00	46.97	99.18	2.55
		26.98	55.43	93.84	2.35
		53.97	82.50	97.09	0.41
Luteoloside	2.92	11.00	13.86	99.42	2.05
		17.46	20.43	100.29	1.60
		34.92	36.98	97.53	0.92
Tricin	0.11	11.00	11.10	99.92	1.21
		17.46	18.66	106.25	1.00
		34.92	35.51	101.37	0.86
Luteolin	0.17	18.00	19.68	108.35	1.55
		28.57	30.13	104.84	1.40
		57.14	62.65	109.33	1.24

Component	BLE1	BLE2	S. tootsik	S. farinosa	S. tenuifolia	B. multiplex	B. chungii	P. edulis	P. maculatus
Orientin	16.74±0.12^c	17.46±0.16^d	4.31±0.10^b	2.23±0.05^a	2.46±0.11^a	-	1.88±0.02^a	38.45±1.17^e	2.27±0.04^a
Isoorientin	34.59±0.41^e	41.14±0.73^f	6.78±0.07^b	4.61±0.08^ab	5.12±0.06^ab	29.85±1.00^d	13.23±0.27^c	101.30±4.82^g	2.71±0.02^a
Vitexin	4.98±0.16^f	4.37±0.17^e	0.67±0.02^b	1.78±0.06^d	4.30±0.28^e	0.99±0.08^c	0.29±0.03^a	9.42±0.37^g	4.23±0.09^e
Isovitexin	11.50±0.04^c	15.44±0.14^d	1.76±0.04^a	1.00±0.06^a	1.27±0.07^a	42.56±1.74^g	19.24±1.01^e	31.14±1.26^f	4.53±0.09^b
Luteoloside	0.36±0.01^a	0.29±0.01^a	8.11±0.17^d	5.35±0.04^c	3.21±0.23^b	15.39±0.26^e	16.76±1.76^f	3.36±0.11^b	17.68±0.26^f
Tricin	0.056±0.001^a	0.052±0.001^a	-	-	-	0.035±0.008^a	0.831±0.018^d	0.103±0.005^b	0.359±0.035^c
Luteolin	0.134±0.003^b	0.790±0.017^g	0.104±0.005^a	-	-	0.363±0.020^f	0.332±0.016^e	0.162±0.006^c	0.189±0.013^d
Total	68.36	79.54	21.73	14.97	16.36	89.19	52.56	183.94	31.97