高效液相色谱-荧光检测法检测烤肉制品中5种硝基多环芳烃

doi:10.3864/j.issn.0578-1752.2021.05.016

摘要/Abstract

摘要：

【目的】明确烤肉制品中硝基多环芳烃（NPAHs）的含量水平,建立固相萃取柱结合高效液相色谱-荧光检测器测定烤肉制品中5种NPAHs（1-硝基萘、2-硝基芴、3-硝基荧蒽、1-硝基芘和6-硝基苯并[a]芘）的方法。【方法】称取1 g真空冷冻干燥的烤肉样品,加入二氯甲烷超声提取并过滤收集滤液,重复提取3次。将滤液氮吹至近干,加入3 mL正己烷溶解。依次用二氯甲烷和正己烷活化多环芳烃固相萃取柱,上样后用正己烷荡洗样品管然后继续上柱;用正己烷淋洗,二氯甲烷洗脱;将洗脱液氮吹至近干,用乙腈溶解得到NPAHs待测液。向待测液中加入酸化甲醇和铁粉,磁力搅拌水浴加热40 min,离心并过滤膜后通过高效液相色谱-荧光检测器检测,其中色谱柱为Agilent ZORBAX Eclipse PAH柱,设置柱温40℃,进样量20 μL,流动相选用水和乙腈,采用梯度洗脱模式。【结果】5种NPAHs在相应质量浓度范围内线性关系良好,相关系数大于0.9940,检出限为0.12—2.17 μg·kg^-1,定量限为0.38—7.23 μg·kg^-1,平均回收率为53.16%—129.64%,精密度为1.91%—30.73%。利用该方法对我国5类典型的烤肉样品进行检测,测得5种NPAHs总含量为50.19—82.36 μg·kg^-1,其中6-硝基苯并[a]芘含量最高,约为31.01—35.89 μg·kg^-1,其次是3-硝基荧蒽,约为9.99—23.06 μg·kg^-1。【结论】固相萃取柱结合高效液相色谱-荧光检测法适用于烤肉制品中NPAHs的分析;我国烤肉制品中普遍存在NPAHs。

关键词: 硝基多环芳烃, 烤肉制品, 高效液相色谱-荧光检测, 固相萃取

Abstract:

【Objective】 This study was aimed to establish an approach to determine the content of five NPAHs (including 1-nitronaphthalene, 2-nitrofluorene, 3-nitrofluoranthene, 1-nitropyrene and 6-nitrobenzo[a]pyrene) in roasted meat based on solid-phase extraction (SPE) column and high performance liquid chromatography with fluorescence detection (HPLC-FLD).【Method】 1 g vacuum freeze-dried roasted meat sample was extracted by dichloromethane for ultrasonic extraction, and the filtrate was collected. Both the extraction and filtration processes were repeated three times. Then, the filtrate was dried by nitrogen and dissolved in 3 mL n-hexane. After that, the extract was loaded onto PAHs SPE column which was previously activated by dichloromethane and n-hexane. After loading, the sample tube was rinsed by n-hexane and loaded as well. The SPE column was washed by n-hexane and eluted by dichloromethane. The eluent was dried by nitrogen, and the residue was dissolved in acetonitrile to obtain NPAHs solution. Acidified methanol and iron powder were added to the NPAHs solution and heated in water bath with magnetic stirring for 40 min. After centrifugation and filtration, 20 μL sample was tested by HPLC-FLD at 40℃ by using Agilent ZORBAX Eclipse PAH analysis column. In the gradient elution system, water and acetonitrile were used as the mobile phases.【Result】 The result showed that five NPAHs were linear with correlation coefficients greater than 0.9940. Specifically, the detection limits (LOD) were within 0.12-2.17 μg·kg^-1, and the quantification limits (LOQ) were within 0.38-7.23 μg·kg ^-1. The average recoveries and the precision were within 53.16%-129.64% and 1.91%-30.73%, respectively. Furthermore, five kinds of typical Chinese roasted meat products were tested by this method, and the content of five NPAHs in all roasted meat samples were between 50.19-82.36 μg·kg ^-1. Among them, the content of 6-nitrobenzo[a]pyrene was the highest, about 31.01-35.89 μg·kg^-1, followed by 3-nitrofluoranthene, and the content was within the range of 9.99-23.06 μg·kg ^-1. 【Conclusion】 SPE column combined with HPLC-FLD was a suitable method to analyze NPAHs in roasted meat products; and NPAHs were widely contained in Chinese roasted meat products.

Key words: nitropolycyclic aromatic hydrocarbon, roasted meat products, high performance liquid chromatography-fluorescence detection, solid-phase extraction

白雪,惠腾,王振宇,曹云刚,张德权. 高效液相色谱-荧光检测法检测烤肉制品中5种硝基多环芳烃[J]. 中国农业科学, 2021, 54(5): 1055-1062.

Xue BAI,Teng HUI,ZhenYu WANG,YunGang CAO,DeQuan ZHANG. Determination of 5 Nitropolycyclic Aromatic Hydrocarbons in Roasted Meat Products by High Performance Liquid Chromatography- Fluorescence Detection[J]. Scientia Agricultura Sinica, 2021, 54(5): 1055-1062.

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链接本文: https://www.chinaagrisci.com/CN/10.3864/j.issn.0578-1752.2021.05.016

https://www.chinaagrisci.com/CN/Y2021/V54/I5/1055

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表5

烤肉制品中NPAHs检测结果"

检测样品 Sample	配料中有无NaNO₂ NaNO₂ in ingredients	1-硝基萘 1-NN (μg?kg^-1)	2-硝基芴 2-NF (μg?kg^-1)	3-硝基荧蒽 3-NF (μg?kg^-1)	1-硝基芘 1-NP (μg?kg^-1)	6-硝基苯并[a]芘 6-NBAP (μg?kg^-1)	总硝基多环芳烃 ΣNPAHs (μg?kg^-1)
北京烤鸭1-胸肉 Beijing roasted duck 1-Breast meat	有 Yes	2.93±0.77b	14.56±4.95a	ND	12.89±0.16bc	31.72±1.29b	70.41±4.18ab
北京烤鸭1-胸皮 Beijing roasted duck 1-Breast skin	有 Yes	3.19±1.21b	12.76±4.52ab	23.06±4.37a	13.80±1.27ab	31.14±0.24b	82.36±8.92a
北京烤鸭1-腿肉 Beijing roasted duck 1-Leg meat	有 Yes	3.41±0.48b	11.71±6.16ab	16.92±1.85abc	12.85±0.08bc	35.89±0.31a	71.18±9.66ab
北京烤鸭1-腿皮 Beijing roasted duck 1-Leg skin	有 Yes	3.64±0.22b	8.91±3.36ab	20.09±7.08abc	13.22±1.09abc	31.83±0.84b	71.92±4.51ab
北京烤鸭2-胸肉 Beijing roasted duck 2-Breast meat	无 No	3.56±0.26b	14.71±0.87a	ND	13.50±1.15abc	31.01±0.17b	77.72±1.65a
北京烤鸭2-胸皮 Beijing roasted duck 2-Breast skin	无 No	3.90±1.02b	10.67±2.56ab	ND	12.75±0.10c	31.02±0.22b	56.39±0.42bc
北京烤鸭2-腿肉 Beijing roasted duck 2-Leg meat	无 No	ND	9.32±3.76ab	ND	12.89±0.14c	31.26±0.46b	58.01±2.78bc
北京烤鸭2-腿皮 Beijing roasted duck 2-Leg skin	无 No	3.81±0.40b	8.33±2.81ab	ND	13.38±0.81abc	31.20±0.32b	55.90±2.50bc
烤羊腿1 Roasted lamb leg 1	有 Yes	3.86±0.69b	9.15±2.86ab	21.16±7.82ab	13.95±0.92a	31.18±0.24b	70.32±7.57ab
烤羊腿2 Roasted lamb leg 2	无 No	10.39±6.07a	11.73±6.68ab	13.35±0.95bc	13.29±0.79abc	31.06±0.21b	64.00±5.81abc
烤猪肉片1 Roasted pork slice 1	有 Yes	3.22±0.26b	7.93±4.94ab	19.80±5.67ab	13.60±0.37abc	31.13±0.28b	68.55±4.88abc
烤猪肉片2 Roasted pork slice 2	无 No	5.77±0.65b	10.33±3.42ab	21.11±5.76ab	12.90±0.16bc	31.07±0.23b	67.73±3.27abc
烤鸡翅1 Roasted chicken wing 1	有 Yes	3.56±0.20b	10.54±3.12ab	9.99±0.99c	12.88±0.26bc	31.29±0.27b	58.63±2.58bc
烤鸡翅2 Roasted chicken wing 2	无 No	5.93±0.20b	8.80±1.31ab	ND	13.06±0.72abc	31.75±0.68b	66.84±2.42abc
烤羊肉串1 Shish kebab 1	无 No	3.91±0.69b	7.25±1.54b	ND	12.95±0.33bc	31.45±1.28b	55.56±1.13bc
烤羊肉串2 Shish kebab 2	无 No	ND	5.84±0.57b	ND	13.22±0.31abc	31.07±0.25b	50.19±0.53c

表5

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硝基多环芳烃 NPAHs	线性方程 Linear equation	相关系数 r	检出限 LOD	定量限 LOQ
1-硝基萘 1-NN	y=1.5533x-0.1532	0.9941	0.20	0.69
2-硝基芴 2-NF	y=1.7404x+0.0286	0.9989	0.22	0.74
3-硝基荧蒽 3-NF	y=0.1666x+0.0218	0.9991	0.79	2.63
1-硝基芘 1-NP	y=2.4321x-3.0797	0.9990	0.12	0.38
6-硝基苯并[a]芘 6-NBAP	y=0.1101x-0.3392	0.9950	2.12	7.23

添加浓度 Added concentration (ng?g^-1)	硝基多环芳烃NPAHs
	1-硝基萘 1-NN		2-硝基芴 2-NF		3-硝基荧蒽 3-NF		1-硝基芘 1-NP		6-硝基苯并[a]芘 6-NBAP
	回收率Recovery (%)	精密度 Precision (%)	回收率Recovery (%)	精密度Precision (%)	回收率Recovery (%)	精密度Precision (%)	回收率Recovery (%)	精密度Precision (%)	回收率Recovery (%)	精密度Precision (%)
20	66.32	1.91	71.62	7.97	82.25	7.75	60.76	2.94	119.60	30.73
60	56.06	4.90	63.43	7.58	71.42	8.48	53.95	5.44	110.20	18.06
100	57.67	5.34	53.16	3.08	75.23	16.30	57.06	5.14	129.64	12.63