超高效液相色谱-串联质谱法同时测定小黄花鱼中9种生物胺

罗振玲; 高海波; 杨挺; 付余

doi:10.13386/j.issn1002-0306.2022040183

超高效液相色谱-串联质谱法同时测定小黄花鱼中9种生物胺

doi: 10.13386/j.issn1002-0306.2022040183

罗振玲^1,,
高海波¹,
杨挺¹,
付余^2, ,

1.
台州市食品检验检测中心，浙江台州 318100
2.
西南大学食品科学学院，重庆 400715

基金项目: 台州市科技计划项目（20ny06）。

详细信息

作者简介:
罗振玲（1989−），女，硕士，工程师，研究方向：食品、农产品质量安全，E-mail：zlluo1989@sina.com

通讯作者:
付余（1987−），男，博士，教授，研究方向：水产食品，E-mail：fuy987@hotmail.com

中图分类号: TS254.1
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出版历程
- 收稿日期: 2022-04-18
- 网络出版日期: 2023-01-12
- 刊出日期: 2023-03-01

Simultaneous Determination of 9 Biogenic Amines in Larimichthys polyactis by UHPLC-MS/MS

LUO Zhenling^1
,,
GAO Haibo¹,
YANG Ting¹,
FU Yu^{2
, ,}

1.
Taizhou Food Inspection Centre, Taizhou 318100, China
2.
College of Food Science, Southwest University, Chongqing 400715, China

摘要

摘要: 为快速检测水产品中精胺、亚精胺、腐胺、尸胺、组胺、色胺、章鱼胺、酪胺、2-苯乙胺9种生物胺含量，本研究以小黄花鱼为基质，优化了流动相组成和样品提取溶剂，建立了超高效液相色谱-串联三重四极杆质谱同时检测方法。样品经0.1%三氯乙酸水溶液重复提取两遍，合并上清液过0.22 μm聚四氟乙烯滤膜。以HSS T3为色谱柱，流动相A为0.1%甲酸乙腈，流动相B为10 mmol/L甲酸铵水溶液含0.2%甲酸，在电喷雾正离子电离、多反应监测模式下，9种生物胺在4 min内得到分离。根据响应强度不同，9种生物胺分成酪胺、2-苯乙胺，腐胺、尸胺、组胺、色胺、章鱼胺，精胺、亚精胺三组，以1:5:25浓度比配制多浓度混合标准中间液。该方法对三组生物胺的线性范围分别为1~40 ng/mL、5~200 ng/mL和25~1000 ng/mL；方法定量限依次为0.01、0.05和0.25 mg/kg。低、中、高三个浓度水平加标回收率分别为81.7%~111.2%、80.0%~110.3%、80.0%~99.0%；精密度（n=6）依次为1.7%~7.3%、1.3%~9.1%、0.7%~4.6%。总体而言，该方法简便、快速，具有良好的灵敏度、回收率和精密度，适用于小黄花鱼中生物胺的测定。
- 生物胺 /
- 超高效液相色谱-串联质谱 /
- 小黄花鱼
Abstract: To rapidly determine 9 biogenic amines including spermine, spermidine, putrescine, cadaverine, histamine, tryptamine, octopamine, tyramine and β-phenethylamine in aquatic products, the ultra high-performance liquid chromatography-tandem mass spectrometry method was established with Larimichthys polyactis using as matrix sample, and mobile phase and extract solvents were optimized. Matrix sample was extracted by 0.1% trichloroacetic acid (TCA) in duplicate, the supernatant was combined and filtered by 0.22 μm PTFE filters, and separated by HSS T3 column in 4 min. The mobile phase A was acetonitrile containing 0.1% formic acid, while mobile phase B was 10 mmol/L ammonium formate containing 0.2% formic acid. Nine biogenic amines were divided into three groups based on their different response intensity, with a concentration range of 1 to 40 ng/mL (tyramine and β-phenethylamine), 5 to 200 ng/mL (putrescine, cadaverine, histamine, tryptamine and octopamine), and 25 to 1000 ng/mL (spermine and spermidine). Their method quantification limits (LOQs) were 0.01, 0.05 and 0.25 mg/kg, respectively. The recovery rate of low, medium and high concentrations was 81.7%~111.2%, 80.0%~110.3% and 80.0%~99.0%, respectively, with the corresponding degree of precision (n=6) of 1.7%~7.3%, 1.3%~9.1% and 0.7%~4.6%. This method is simple, fast and with good sensitivity, recovery and precision, which is suitable for the determination of biogenic amines in Larimichthys polyactis.
- biogenic amines /
- ultra high-performance liquid chromatography-tandem mass spectrometry (UHPLC-MS/MS) /
- Larimichthys polyactis

HTML全文

图 1 九种不同生物胺在四组流动相中的色谱图

Figure 1. Total ion chromatogram of 9 BAs standards in 4 group mobile phases

注：组胺、2-苯乙胺及色胺绘在次纵坐标上。

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表 1 流动相梯度洗脱程序

Table 1. The linear gradient program of mobile phase

时间（min）	流动相A（%）	流动相B（%）
0	5	95
0.3	5	95
4.5	50	50
5.0	95	5
6.5	95	5
6.6	5	95
8.0	5	95

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表 2 九种生物胺多反应监测扫描模式的质谱参数

Table 2. Parameters of MS/MS for 9 biogenic amines (BAs) MRM mode

化合物	质荷比（m/z）		去簇电压（V）	碰撞电压（V）
化合物	母离子	子离子	去簇电压（V）	碰撞电压（V）
精胺	203.2	129.1^* 112.1	55 120	25 47
亚精胺	146.0	129.1^* 112.1	40 40	16 19
腐胺	89.0	72.0^* 30.0	50 50	15 15
尸胺	103.2	86.0^* 69.1	43 50	14 21
组胺	112.2	94.7^* 68.1	71 70	25 29
色胺	161.1	144.1^* 117.1	40 41	16 35
章鱼胺	154.0	136.2^* 91.1	45 31	11 29
酪胺	138.0	77.0^* 93.0	43 42	43 24
2-苯乙胺	122.0	105.2^* 79.1	41 42	19 31
注：*为定量离子。

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表 3 不同提取溶剂中小黄花鱼生物胺加标回收率与精密度（n=6）

Table 3. Recoveries and precision of 9 BAs spiked in Larimichthys polyactis by different extract solvents (n=6)

化合物	添加量（mg/kg）	0.05% TCA水溶液			0.1% TCA水溶液			0.1% TCA水溶液:乙腈（9:1，v/v）
化合物	添加量（mg/kg）	本底值（mg/kg）	回收率（%）	RSD（%）	本底值（mg/kg）	回收率（%）	RSD（%）	本底值（mg/kg）	回收率（%）	RSD（%）
精胺	8 15 40	0.23±0.03^a	85.6 87.8 79.7	3.5 3.2 1.8	1.77±0.05^c	106.2 103.8 94.3	1.7 9.1 1.1	1.52±0.05^b	74.6 82.5 99.8	1.8 1.6 1.4
亚精胺	8 15 40	0.35±0.01^a	62.8 79.7 91.0	2.8 2.5 2.0	0.74±0.03^c	111.2 110.3 99.0	3.6 3.8 1.3	0.68±0.03^b	89.5 113.1 90.2	6.7 1.7 1.2
腐胺	1.6 3.0 8.0	1.15±0.03^b	107.4 91.9 73.3	1.3 1.4 0.9	1.38±0.08^c	103.2 98.9 80.7	6.6 1.3 2.9	1.02±0.04^a	102.7 92.4 75.2	3.1 3.3 0.8
尸胺	1.6 3.0 8.0		78.8 72.0 72.9	5.3 1.1 1.1		90.3 83.7 80.4	7.3 5.7 1.6		80.7 73.3 72.3	6.0 4.1 1.9
组胺	1.6 3.0 8.0		79.0 62.8 60.2	2.3 5.2 1.4		87.6 88.0 87.4	3.3 4.7 1.3		85.6 73.3 72.3	2.8 4.1 0.7
色胺	1.6 3.0 8.0		85.4 80.2 64.5	5.3 4.8 2.5		88.1 80.5 80.2	5.7 7.7 4.6		89.2 78.8 75.2	3.6 1.4 1.7
章鱼胺	1.6 3.0 8.0		57.5 56.1 59.9	8.6 4.5 1.1		85.3 80.5 80.4	5.6 7.0 1.6		70.0 70.2 70.2	5.9 1.1 1.2
酪胺	0.32 0.60 1.60		66.8 67.2 64.2	5.4 4.2 2.2		81.7 80.0 80.0	5.9 2.4 0.7		75.2 62.5 70.1	2.9 4.4 1.0
2-苯乙胺	0.32 0.60 1.60		91.3 86.4 90.3	6.4 5.4 0.9		92.4 90.4 91.3	4.3 4.1 2.2		89.7 91.2 89.9	5.2 2.5 0.7
注：不同提取溶剂对小黄花鱼中同种生物胺本底的检测结果采用Duncan’s 多重比较；同行不同小写字母表示分析结果差异显著（P<0.05）。

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表 4 九种生物胺的线性范围、回归方程、相关系数（r）、检出限和定量限

Table 4. Linear range, calibration curves, correlation coefficient (r), limit of detection (LOD) and limit of quantitation (LOQ) of 9 BAs

化合物	线性范围（ng/mL）	回归方程	相关系数r	检出限（mg/kg）	定量限（mg/kg）
精胺	25~1000	y=3317.9x−25504	0.9992	0.05	0.25
亚精胺	25~1000	y=2396.4x+45234	0.9988	0.05	0.25
腐胺	5~200	y=4902.3x+265395	0.9956	0.01	0.05
尸胺	5~200	y=8235x+116707	0.9954	0.01	0.05
组胺	5~200	y=67903x+530114	0.9974	0.01	0.05
色胺	5~200	y=56051x+234899	0.9987	0.01	0.05
章鱼胺	5~200	y=5197.7x+22794	0.9988	0.01	0.05
酪胺	1~40	y=22708x+1850.3	0.9999	0.003	0.01
2-苯乙胺	1~40	y=219501x+5405.5	0.9999	0.003	0.01

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表 5 不同水产品中9种生物胺含量的测定结果（mg/kg, n=6）

Table 5. The detection results of 9 BAs in different aquatic products (mg/kg, n=6)

样品	精胺	亚精胺	腐胺	尸胺	组胺	色胺	章鱼胺	酪胺	2-苯乙胺
小黄花鱼I	1.64±0.14	0.67±0.04	1.26±0.05	/	/	/	/	/	/
小黄花鱼II	1.79±0.05	0.63±0.11	1.38±0.21	/	/	/	/	/	/
小黄花鱼III	1.54±0.19	0.52±0.03	1.42±0.16	/	/	/	/	/	/
小黄花鱼IV	1.92±0.31	0.72±0.15	1.55±0.19	/	/	/	/	/	/
小黄花鱼V	1.47±0.22	0.65±0.02	1.48±0.09	/	/	/	/	/	/
水潺	2.02±0.21	3.21±0.24	3.69±0.12	/	/	/	/	/	/
鲳鱼	1.18±0.05	0.85±0.03	3.27±0.02	0.20±0.03	/	/	/	0.15±0.14	/
带鱼	1.06±0.08	0.48±0.05	0.84±0.05	/	/	/	/	/	/
墨鱼	7.64±0.19	1.81±0.14	1.90±0.16	3.07±0.14	/	/	/	/	/
鱿鱼	21.3±3.07	4.06±0.30	1.49±0.17	1.58±0.10	/	/	/	/	/
红虾	0.73±0.03	0.64±0.00	/	0.34±0.01	/	/	/	0.65±0.00	0.26±0.01
南美白对虾	0.69±0.02	0.76±0.00	/	0.57±0.01	/	/	/	0.75±0.04	/
死后3~5 h南美白对虾	0.71±0.01	3.16±0.36	2.34±0.17	9.80±0.16	/	/	/	1.01±0.03	/
注：“/”表示未检出。

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