气相色谱-氢火焰离子化检测器测定婴幼儿乳粉中37种脂肪酸含量

罗叶丽; 李国薇; 董梅; 赵璧; 强鑫

doi:10.13386/j.issn1002-0306.2022040281

气相色谱-氢火焰离子化检测器测定婴幼儿乳粉中37种脂肪酸含量

doi: 10.13386/j.issn1002-0306.2022040281

罗叶丽^1,,
李国薇^2, ,,
董梅¹,
赵璧²,
强鑫²

1.
渭南市检验检测研究院，陕西渭南 714000
2.
富平县检验检测中心（陕西省羊乳产品质量监督检验中心），陕西富平 711700

详细信息

作者简介:
罗叶丽（1988−），女，硕士，中级工程师，研究方向：食品营养物质、重金属及有毒有害物质的检测，E-mail：syh_200@126.com

通讯作者:
李国薇（1988−），女，硕士，中级工程师，研究方向：食品营养物质及有害成分检测，E-mail：xi_aoweilove_ok@126.com

中图分类号: O657.7⁺1
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出版历程
- 收稿日期: 2022-04-25
- 网络出版日期: 2023-05-06
- 刊出日期: 2023-06-01

Determination of 37 Kinds of Fatty Acids in Infant Milk Powder by Gas Chromatography-Hydrogen Flame Ionization Detector

LUO Yeli^1
,,
LI Guowei^{2
, ,},
DONG Mei¹,
ZHAO Bi²,
QIANG Xin²

1.
Weinan Inspection and Research Institute, Weinan 714000, China
2.
Fuping County Inspection and Testing Center (Shaanxi Sheep Milk Product Quality Supervision and Inspection Center), Fuping 711700, China

摘要

摘要: 目的：建立气相色谱-氢火焰离子化检测器测定乳粉中37种脂肪酸含量的分析方法。方法：婴幼儿乳粉样品运用乙酰氯-甲醇法（10%乙酰氯甲醇溶液于80 ℃±1 ℃水浴2 h）甲酯化，离心取上清液过0.22 μm的有机滤膜后经SP-2560（100 m×250 μm×0.2 μm）分离，气相色谱法检测，外标法定量。结果：37种脂肪酸甲酯在0.002~0.2 mg/mL浓度范围内线性良好，相关系数均大于0.999，回收率低梯度在85.5%~100.0%之间，中梯度在91.2%~104.2%之间，高梯度在86.8%~103.8%之间，精密度在0.88%~3.81%之间，实际样品测定结果符合相关标准要求。结论：该方法是在国标基础上建立了一种更为快速、分离度更好的脂肪酸检测技术，有很好的实用价值，方法稳定性好，成本低，重现性好，准确度高，适用于婴幼儿乳粉中37种脂肪酸含量的测定。
- 脂肪酸 /
- 乙酰氯-甲醇法 /
- 气相色谱法 /
- 婴幼儿乳粉
Abstract: Objective: To establish an analytical method for the determination of 37 kinds of fatty acid in milk powder by gas chromatography-hydrogen flame ionization detector (GC-FID). Methods: Infant milk powder samples were methylated by acetyl chloride-methanol method (10% acetyl chloride methanol solution in water bath at 80 ℃±1 ℃ for 2 h). The supernatant was collected by centrifugation, passed through a 0.22 μm organic filter membrane, and separated through SP-2560 (100 m×250 μm×0.2 μm). Then, the detection was operated by gas chromatography, and the external standard method was used for quantification. Results: The linearity of 37 kinds of fatty acid methyl esters was good in the concentration range of 0.002~0.2 mg/mL, and the correlation coefficients were all greater than 0.999. The recovery rate fell in 85.5%~100.0% for the low gradient, 91.2%~104.2% for the medium gradient, and 86.8%~103.8% for the high gradient. The precision was between 0.88% and 3.81%, and the actual sample determination results met the relevant standards. Conclusion: This method is a more rapid and better separation degree of fatty acid detection technology established on the basis of national standard, which has good practical value. The method has good stability, low cost, good reproducibility and high accuracy, and it is suitable for the determination of 37 kinds of fatty acid in infant milk powder.
- fatty acid /
- acetyl chloride-methanol method /
- gas chromatography /
- infant milk powder

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图 1 37种脂肪酸甲酯色谱图

Figure 1. The chromatomap of 37 kinds fatty acid methyl esters

注：1.丁酸甲酯（C4:0），2.己酸甲酯（C6:0），3.辛酸甲酯（C8:0），4.癸酸甲酯（C10:0），5.十一碳酸甲酯（C11:0），6.月桂酸甲酯（C12:0），7.十三碳酸甲酯（C13:0），8.肉豆蔻酸甲酯（C14:0），9.肉豆蔻烯酸甲脂（C14:1），10.十五碳酸甲脂（C15:0），11.十五碳一烯酸甲酯（C15:1），12.棕榈酸甲酯（C16:0），13.十六碳烯酸甲酯（C16:1），14.十七碳酸甲脂（C17:0），15.十七碳一烯酸甲酯（C17:1），16.硬脂酸甲酯（C18:0），17.反式油酸甲酯（C18:1n9t），18.油酸甲酯（C18:1n9c），19.反式亚油酸甲酯（C18:2n6t），20.亚油酸甲酯（C18:2n6c），21.花生酸甲酯（C20:0），22.亚麻酸甲酯（C18:3n6），23.二十碳烯酸甲酯（C20:1），24.二十一碳酸甲酯（C21:0），25.α-亚麻酸甲酯（C18:3n3），26.二十碳二烯酸甲酯（C20:2），27.二十二碳酸甲酯（C22:0），28.二十碳三烯酸甲酯（C20:3n6），29.芥酸甲酯（C22:1n9），30.二十碳三烯酸甲酯（C20:3n3），31.花生四烯酸 ARA甲酯（C20:4n6），32.二十三碳酸甲酯（C23:0），33.二十二碳二烯酸甲酯（C22:2），34.二十四碳酸甲酯（C24:0），35.二十碳五烯酸EPA甲酯（C20:5n3），36.二十四碳一烯酸甲酯（C24:1），37.二十二碳六烯酸甲酯DHA甲酯（C22:6n3）。

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图 2 37种脂肪酸甲酯色谱图（GB 5009.168-2016）

Figure 2. The chromatomap of 37 kinds fatty acid methyl esters (GB 5009.168-2016)

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图 3 37种脂肪酸甲酯色谱图（某乳粉企业）

Figure 3. The chromatomap of 37 kinds fatty acid methyl esters (A milk powder enterprise)

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4 实际样品测定代表谱图

4. The representative chromatomap of actual samples

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表 1 色谱条件优化结果统计

Table 1. Statistics of the optimized results of the chromatographic conditions

序号	程序升温	流速（mL/min）	结果	举例
1	130 ℃保持 5 min， 4 ℃/min 升至240 ℃，保持30 min	0.40	峰总数36个
2		0.45	峰总数37个，分离度满足要求	如图1
3		0.50	峰总数37个，有双头峰出现，分离度不满足要求
4		0.55	峰总数37个，有双头峰出现，分离度不满足要求
5		0.60	峰总数36个
6	130 ℃保持 5 min， 3 ℃ /min升至240 ℃，保持30 min	0.40	峰总数36个
7		0.45	峰总数36个，有双头峰出现
8		0.50	峰总数35个

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表 2 色谱峰定性分析结果

Table 2. Results of the qualitative analysis of the chromatographic peaks

序号	名称	单标时间（min）	混标时间（min）	序号	名称	单标时间（min）	混标时间（min）	序号	名称	单标时间（min）	混标时间（min）
1	C4:0	18.74	18.684	14	C17:0	36.513	36.442	27	C22:0	45.705	45.597
2	C6:0	19.863	19.808	15	C17:1	37.938	37.869	28	C20:3n6	47.126	47.080
3	C8:0	21.697	21.646	16	C18:0	38.261	38.172	29	C22:1n9	47.340	47.262
4	C10:0	24.388	24.336	17	C18:1n9t	39.142	39.071	30	C20:3n3	48.036	47.756
5	C11:0	26.001	25.947	18	C18:1n9c	39.556	39.489	31	C20:4n6	48.592	47.997
6	C12:0	27.721	27.667	19	C18:2n6t	40.502	40.436	32	C23:0	47.856	48.556
7	C13:0	29.493	29.438	20	C18:2n6c	41.475	41.413	33	C22:2	49.862	49.810
8	C14:0	31.273	31.214	21	C20:0	41.832	41.731	34	C24:0	50.229	50.108
9	C14:1	32.873	32.819	22	C18:3n6	42.997	42.940	35	C20:5n3	51.847	51.816
10	C15:0	33.026	32.962	23	C20:1	43.229	43.159	36	C24:1	52.248	52.152
11	C15:1	34.639	34.585	24	C18:3n3	43.802	43.608	37	C22:6n3	60.507	60.458
12	C16:0	34.768	34.703	25	C21:0	43.708	43.748
13	C16:1	36.163	36.108	26	C20:2	45.369	45.314

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表 3 37种脂肪酸甲酯的线性回归方程

Table 3. The linear equations of the 37 kinds of fatty acid methyl esters

序号	化合物	浓度范围（mg/mL）	回归方程	相关系数r	序号	化合物	浓度范围（mg/mL）	回归方程	相关系数r
1	C4:0	0.002~0.2	Y=287.47101X+0.265797	0.99984	20	C18:2n6c	0.002~0.2	Y=495.43415X−0.0999114	0.99997
2	C6:0	0.002~0.2	Y=369.98152X+0.0486260	0.99998	21	C20:0	0.002~0.2	Y=222.91650X−0.233088	0.99959
3	C8:0	0.002~0.2	Y=395.21120X−0.0443436	0.99998	22	C18:3n6	0.002~0.2	Y=462.77099X−0.211804	0.99992
4	C10:0	0.002~0.2	Y=444.72122X−0.0516759	0.99996	23	C20:1	0.002~0.2	Y=485.68875X−0.229321	0.99991
5	C11:0	0.002~0.2	Y=411.03663X−0.0924383	0.99996	24	C21:0	0.002~0.2	Y=254.94246X−0.135998	0.99985
6	C12:0	0.002~0.2	Y=455.26881X−0.0524545	0.99996	25	C18:3n3	0.002~0.2	Y=567.76917X−0.215109	0.99994
7	C13:0	0.002~0.2	Y=481.55945X−0.136193	0.99996	26	C20:2	0.002~0.2	Y=490.44683X−0.0988460	0.99996
8	C14:0	0.002~0.2	Y=476.33386X−0.0509038	0.99998	27	C22:0	0.002~0.2	Y=245.07514X−0.0848123	0.99994
9	C14:1	0.002~0.2	Y=413.36235X−0.148176	0.99995	28	C20:3n6	0.002~0.2	Y=478.73660X−0.0978376	0.99996
10	C15:0	0.002~0.2	Y=438.10240X−0.200717	0.99993	29	C22:1n9	0.002~0.2	Y=505.07453X+0.00328606	0.99998
11	C15:1	0.002~0.2	Y=472.20237X−0.154981	0.99996	30	C20:3n3	0.002~0.2	Y=375.94478X−0.342025	0.99972
12	C16:0	0.002~0.2	Y=465.70415X−0.0167539	9.99999	31	C20:4n6	0.002~0.2	Y=472.57919X−0.161708	0.99991
13	C16:1	0.002~0.2	Y=476.21573X−0.0970916	0.99998	32	C23:0	0.002~0.2	Y=450.75622X−0.0716064	0.99993
14	C17:0	0.002~0.2	Y=495.77014X−0.117625	0.99997	33	C22:2	0.002~0.2	Y=471.87786X−0.192971	0.99991
15	C17:1	0.002~0.2	Y=525.93097X−0.153926	0.99996	34	C24:0	0.002~0.2	Y=300.61982X+0.00606836	0.99996
16	C18:0	0.002~0.2	Y=190.11571X+0.0611815	0.99999	35	C20:5n3	0.002~0.2	Y=426.16295X−0.0741455	0.99993
17	C18:1n9t	0.002~0.2	Y=480.50965X−0.154013	0.99992	36	C24:1	0.002~0.2	Y=503.25603X−0.00719366	0.99990
18	C18:1n9c	0.002~0.2	Y=503.07903X−0.0732298	0.99998	37	C22:6n3	0.002~0.2	Y=425.76968X−0.321497	0.99991
19	C18:2n6t	0.002~0.2	Y=478.81231X−0.0617369	0.99997

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表 4 定量限验证结果

Table 4. The validation results of LOQ

序号	化合物	理论定量限（g/100 g）	测得含量（g/100 g）	信噪比	序号	化合物	理论定量限（g/100 g）	测得含量（g/100 g）	信噪比
1	C4:0	0.0033	0.0040	13.0	20	C18:2n6c	0.0033	0.0037	24.6
2	C6:0	0.0033	0.0039	16.4	21	C20:0	0.0066	0.0079	13.0
3	C8:0	0.0033	0.0036	17.8	22	C18:3n6	0.0066	0.0070	23.8
4	C10:0	0.0066	0.0073	22.8	23	C20:1	0.0033	0.0037	23.7
5	C11:0	0.0033	0.0031	18.4	24	C21:0	0.0033	0.0035	12.3
6	C12:0	0.0066	0.0073	25.1	25	C18:3n3	0.0033	0.0036	25.8
7	C13:0	0.0033	0.0035	25.3	26	C20:2	0.0033	0.0035	21.8
8	C14:0	0.0033	0.0034	25.4	27	C22:0	0.0066	0.0051	10.4
9	C14:1	0.0033	0.0036	23.9	28	C20:3n6	0.0033	0.0031	18.3
10	C15:0	0.0033	0.0037	26.0	29	C22:1n9	0.0033	0.0033	21.3
11	C15:1	0.0033	0.0035	25.6	30	C20:3n3	0.0033	0.0035	16.2
12	C16:0	0.0066	0.0071	28.7	31	C20:4n6	0.0033	0.0034	18.8
13	C16:1	0.0033	0.0037	24.7	32	C23:0	0.0033	0.0035	17.5
14	C17:0	0.0066	0.0070	29.9	33	C22:2	0.0033	0.0036	17.8
15	C17:1	0.0033	0.0034	27.5	34	C24:0	0.0066	0.0070	13.0
16	C18:0	0.0066	0.0076	11.4	35	C20:5n3	0.0033	0.0036	14.9
17	C18:1n9t	0.0033	0.0034	24.1	36	C24:1	0.0033	0.0034	17.6
18	C18:1n9c	0.0066	0.0072	28.6	37	C22:6n3	0.0033	0.0025	10.2
19	C18:2n6t	0.0033	0.0033	23.1

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表 5 方法精密度实验结果（n=6）

Table 5. Results of precision for this method (n=6)

序号	名称	精密度（%）	序号	名称	精密度（%）	序号	名称	精密度（%）	序号	名称	精密度（%）
1	C4:0	2.29	11	C15:1	1.23	21	C20:0	3.05	31	C20:4n6	1.47
2	C6:0	1.81	12	C16:0	1.83	22	C18:3n6	2.01	32	C23:0	1.87
3	C8:0	1.59	13	C16:1	1.86	23	C20:1	0.93	33	C22:2	1.28
4	C10:0	1.87	14	C17:0	1.57	24	C18:3n3	1.97	34	C24:0	1.17
5	C11:0	1.85	15	C17:1	1.90	25	C21:0	1.75	35	C20:5n3	2.44
6	C12:0	1.82	16	C18:0	3.71	26	C20:2	2.16	36	C24:1	2.19
7	C13:0	2.14	17	C18:1n9t	2.35	27	C22:0	3.81	37	C22:6n3	1.47
8	C14:0	1.67	18	C18:1n9c	1.45	28	C20:3n6	1.29
9	C14:1	0.88	19	C18:2n6t	2.30	29	C22:1n9	1.82
10	C15:0	1.19	20	C18:2n6c	1.73	30	C20:3n3	1.84

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表 6 回收率实验结果

Table 6. Results of recoveries for this method

序号	检测项目	本底值（g/100 g）	理论加标量（g/100 g）			实测加标量（g/100 g）			回收率（%）
序号	检测项目	本底值（g/100 g）	低	中	高	低	中	高	低	中	高
1	C11:0	0.0344	0.0055	0.0175	0.0332	0.0398	0.0515	0.0674	98.2	97.7	99.4
2	C13:0	0.0198	0.0065	0.0186	0.0372	0.0261	0.0383	0.0521	96.9	99.5	86.8
3	C14:1	0.0196	0.0084	0.0224	0.0402	0.0271	0.0412	0.0611	89.3	96.4	103.2
4	C15:0	0.091	0.0075	0.0188	0.0328	0.0975	0.110	0.123	86.7	101.1	97.6
5	C15:1	0.00336	0.0075	0.0187	0.0328	0.0102	0.0211	0.0374	91.2	94.9	103.8
6	C16:1	0.0653	0.0047	0.0150	0.0301	0.0699	0.0809	0.0943	97.9	104.0	96.3
7	C17:0	0.056	0.0057	0.0170	0.0302	0.0612	0.0721	0.0860	91.2	94.7	99.3
8	C18:2n6t	0.00934	0.0076	0.0208	0.0397	0.0163	0.0291	0.0488	91.6	95.0	99.4
9	C18:3n6	0.0207	0.0076	0.0189	0.0397	0.0272	0.0395	0.0581	85.5	99.5	94.2
10	C20:1	0.0302	0.0066	0.0190	0.0361	0.0367	0.0494	0.0672	98.5	101.1	102.5
11	C21:0	0.0124	0.0076	0.0190	0.0399	0.0193	0.0316	0.0501	90.8	101.1	94.5
12	C20:2	0.00839	0.0085	0.0209	0.0398	0.0162	0.0291	0.0475	91.9	99.1	98.3
13	C22:0	0.0758	0.0048	0.0152	0.0286	0.0803	0.0905	0.103	93.7	96.7	95.1
14	C20:3n6	0.0139	0.0076	0.0190	0.0398	0.0211	0.0337	0.0531	94.7	104.2	98.5
15	C20:3n3	0.0185	0.0066	0.0190	0.0398	0.0243	0.0372	0.0567	87.9	98.4	96.0
16	C23:0	0.0819	0.0076	0.0143	0.0296	0.0891	0.0962	0.110	94.7	100.0	94.9
17	C22:2	0.00601	0.0076	0.0209	0.0394	0.0132	0.0260	0.0464	94.6	95.6	102.5
18	C24:0	0.0525	0.0057	0.0162	0.0325	0.0582	0.0686	0.0841	100.0	99.4	97.2
19	C24:1	0.00808	0.0076	0.0191	0.0363	0.0146	0.0255	0.0431	85.8	91.2	96.5

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表 7 实际样品测定结果

Table 7. Results of actual samples

序号	检测项目	一段（g/100 g）	二段-1 （g/100 g）	二段-2 （g/100 g）	三段（g/100 g）	序号	检测项目	一段（g/100 g）	二段-1 （g/100 g）	二段-2 （g/100 g）	三段（g/100 g）
1	C4:0	0.144	0.359	0.391	0.349	20	C18:2n6c	3.15	1.96	2.36	1.85
2	C6:0	0.169	0.223	0.260	0.211	21	C20:0	0.187	0.122	0.139	0.115
3	C8:0	0.443	0.246	0.282	0.241	22	C18:3n6	0.0171	0.0196	0.0115	0.0207
4	C10:0	0.716	0.762	0.852	0.741	23	C20:1	0.0324	0.0293	0.0265	0.0302
5	C11:0	0.0192	0.0337	0.0326	0.0344	24	C21:0	0.0398	0.0104	0.0114	0.0124
6	C12:0	1.41	0.417	0.429	0.408	25	C18:3n3	0.397	0.172	0.241	0.165
7	C13:0	0.0105	0.0183	0.00743	0.0198	26	C20:2	0.00385	0.00608	0.00343	0.00839
8	C14:0	1.16	0.856	0.915	0.830	27	C22:0	0.461	0.0783	0.0966	0.0758
9	C14:1	0.0111	0.0179	0.0171	0.0196	28	C20:3n6	0.00973	0.0128	0.00409	0.0139
10	C15:0	0.0603	0.0952	0.106	0.0910	29	C22:1n9	0.00176	0.00558	0.00230	0.00818
11	C15:1	0.0155	0.0227	0.000683	0.00336	30	C20:3n3	0.0390	0.0156	0.0172	0.0185
12	C16:0	4.81	4.63	4.74	4.40	31	C20:4n6	0.00217	0.00504	0.00193	0.00747
13	C16:1	0.0481	0.0668	0.0733	0.0653	32	C23:0	0.105	0.0911	0.0237	0.0819
14	C17:0	0.0481	0.0566	0.0679	0.0560	33	C22:2	0.00155	0.00354	0.00241	0.00601
15	C17:1	0.0226	0.0300	0.0323	0.0315	34	C24:0	0.181	0.0548	0.0334	0.0525
16	C18:0	1.15	2.90	3.45	2.69	35	C20:5n3	0.00458	0.00851	0.00681	0.00981
17	C18:1n9t	0.187	0.190	0.243	0.186	36	C24:1	0.00319	0.00698	0.00254	0.00808
18	C18:1n9c	4.50	4.62	4.88	4.41	37	C22:6n3	0.0609	0.0494	0.00378	0.0455
19	C18:2n6t	0.00285	0.00673	0.00401	0.00934
亚油酸:α-亚麻酸		8:1	11:1	10:1	11:1	总脂肪酸（g/100 g）		19.6	18.2	19.8	17.3
DHA/%总脂肪酸		0.310	0.271	0.0191	0.263	（月桂酸+肉豆蔻酸）/%总脂肪酸		13.1	6.99	6.80	7.15
ARA/%总脂肪酸		0.0111	0.0277	0.00976	0.0431	芥酸/%总脂肪酸		0.00897	0.430	0.0207	0.0472
反式脂肪酸/%总脂肪酸		0.967	1.08	1.25	1.13

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