Simultaneous determination of 37 mycotoxins in grain and animal feed by impurity adsorption purification coupled with ultra-performance liquid chromatography-tandem mass spectrometry

doi:10.3724/SP.J.1123.2019.12013

Abstract

Abstract:

A rapid method based on ultra-performance liquid chromatography-tandem mass spectrometry (UPLC–MS/MS) was developed for the simultaneous determination of 37 mycotoxins having various physicochemical properties in grain and animal feed samples. The 37 analytes were extracted from ground samples with acetonitrile-water-formic acid (84:15.9:0.1, v/v/v) by 20 min vibration, and purified using a commercial MLJ-1 pass-through solid-phase extraction (SPE) clean-up cartridge. The analytes were then separated on a reversed-phase BEH RP18 column by a gradient elution program with 0.1 mmol/L ammonium acetate (containing 0.1% (v/v) formic acid) aqueous solution and 0.1% (v/v) formic acid methanol solution as mobile phases. The separated analytes were detected by MS/MS in the multiple reaction monitoring (MRM) mode via ESI+/- ionization. The results showed that the purification was completed in 1 min and that the 37 analytes could be separated on the chromatographic column in 15 min. The 37 mycotoxins showed a linear relationship within their respective linear ranges, and the correlation coefficients of the matrix-matched calibration curves were greater than 0.98. The average recoveries at four spiked levels (limit of quantification (LOQ), LOQ×5, LOQ×10, LOQ×25) for all the targets except fumonisins ranged from 80% to 120%, with the relative standard deviations (RSDs) lower than 20% (n=6). The limits of quantification (LOQs) for all the analytes were between 2 and 40 μg/kg. The proposed method is simple, fast, and accurate, thus being suitable for detecting multiple mycotoxins in grain and animal feed samples.

Key words: impurity adsorption, solid phase extraction (SPE), ultra-performance liquid chromatography-tandem mass spectrometry (UPLC-MS/MS), mycotoxins, grain, feed

WANG Ruiguo, GUO Lili, WANG Peilong, SU Xiao'ou, SONG Zhichao, LIN Gang, ZHU Ronghua. Simultaneous determination of 37 mycotoxins in grain and animal feed by impurity adsorption purification coupled with ultra-performance liquid chromatography-tandem mass spectrometry[J]. Chinese Journal of Chromatography, 2020, 38(7): 817-825.

Figures/Tables 7

References

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Group	No.	Compound	Retention time/min	Adduct	Precursor ion (m/z)	Product ions (m/z)	CV/V	CE/eV
*Quantification ion.
A	1	aflatoxin B₁ (黄曲霉毒素B₁)	8.25	[M+H]⁺	313.1	284.9^*, 241.0	46	26, 40
	2	aflatoxin B₂ (黄曲霉毒素B₂)	7.92	[M+H]⁺	315.2	287.1^*, 259.0	46	30, 32
	3	aflatoxin G₁ (黄曲霉毒素G₁)	7.73	[M+H]⁺	329.1	242.9^*, 200.0	42	28, 42
	4	aflatoxin G₂ (黄曲霉毒素G₂)	7.39	[M+H]⁺	331.1	189.0^*, 245.0	48	43, 34
	5	sterigmatocystin (杂色曲霉素)	10.81	[M+H]⁺	324.8	310.0^*, 280.8	45	26, 46
B	6	ochratoxin A (赭曲霉毒素A)	10.61	[M+H]⁺	404.1	238.9^*, 220.9	32	28, 40
	7	ochratoxin B (赭曲霉毒素B)	9.83	[M+H]⁺	370.2	204.9^*, 102.8	25	25, 54
	8	T-2 toxin (T-2毒素)	9.63	[M+NH₄]⁺	484.2	185.1^*, 305.3	24	25, 18
	9	T-2 triol (T-2三醇)	8.34	[M+Na]⁺	405.2	303.1^*, 125.0	62	14, 16
	10	HT-2 toxin (HT-2毒素)	9.03	[M+NH₄]⁺	442.2	263.2^*, 245.1	30	10, 10
	11	lysergol (麦角醇)	4.32	[M+H]+	254.8	197.0^*, 44.0	15	24, 15
	12	ergocornine (麦角柯宁碱)	7.72	[M+H]⁺	562.2	208.0^*, 223.1	36	48, 40
	13	zearalenone (玉米赤霉烯酮)	10.83	[M-H]^-	317.2	131.0^*, 175.1	48	35, 30
	14	α-zearalenol (α-玉米赤霉烯醇)	10.96	[M-H]^-	319.0	275.1^*, 301.1	44	33, 30
	15	β-zearalenol (β-玉米赤霉烯醇)	10.34	[M-H]^-	319.0	275.1^*, 301.1	42	32, 28
	16	zearalanone (玉米赤霉酮)	10.70	[M-H]^-	319.0	275.1^*, 301.1	43	28, 34
	17	α-zearalanol (α-玉米赤霉醇)	10.69	[M-H]^-	321.0	277.2^*, 303.2	42	26, 26
	18	β-zearalanol (β-玉米赤霉醇)	10.11	[M-H]^-	321.0	277.2^*, 303.2	40	28, 32
C	19	deoxynivalenol (脱氧雪腐镰刀菌烯醇)	4.38	[M+H]⁺	296.9	249.1^*, 231.1	15	10, 15
	20	15-acetyldeoxynivalenol	6.75	[M+H]⁺	338.6	137.0^*, 321.0	25	22, 15
		(15-乙酰基脱氧雪腐镰刀菌烯醇)
	21	3-acetyldeoxynivalenol	6.75	[M+H]⁺	339.0	231.0^*, 203.1	28	16, 22
		(3-乙酰基脱氧雪腐镰刀菌烯醇)
	22	fumonisin B₁ (伏马毒素B₁)	8.69	[M+H]⁺	722.2	352.3^*, 334.3	50	36, 40
	23	fumonisin B₂ (伏马毒素B₂)	9.49	[M+H]⁺	706.4	336.5^*, 354.5	40	40, 35
	24	fumonisin B₃ (伏马毒素B₃)	9.14	[M+H]⁺	706.4	336.5^*, 354.5	40	40, 34
	25	fusarenon X (镰刀菌烯酮-X)	5.65	[M+H]⁺	355.1	247.1^*, 175.1	24	14, 22
	26	diacetoxyscirpenol (蛇形菌素)	8.07	[M+NH₄]⁺	384.2	307.2^*, 105.0	25	15, 30
	27	neosolaniol (新茄病镰刀菌烯醇)	6.09	[M+NH₄]⁺	400.2	185.0^*, 215.1	24	25, 20
	28	tentoxin (腾毒素)	9.32	[M+H]⁺	415.2	312.2^*, 256.1	66	20, 28
	29	patulin (展青霉素)	2.30	[M+H]⁺	155.0	71.0^*, 81.0	30	10, 10
	30	glotoxin (胶霉毒素)	8.30	[M+H]⁺	327.4	263.1^*, 245.1	26	14, 22
	31	verruculogen (疣孢青霉原)	10.71	[M+Na]⁺	534.0	392.1^*, 435.2	30	18, 20
	32	penitrem A (青霉震颤素A)	11.78	[M+H]⁺	634.3	558.3^*, 616.3	22	18, 10
	33	penicillic acid (青霉酸)	5.46	[M+H]⁺	171.0	97.0^*, 82.0	20	17, 25
	34	mycophenolic acid (霉酚酸)	9.69	[M+H]⁺	320.9	207.0^*, 159.0	25	26, 32
	35	roquefortine C (娄地青霉素C)	7.89	[M+H]⁺	389.9	193.0^*, 108.2	40	30, 42
	36	citreoviridin (黄绿青霉素)	10.38	[M+H]⁺	403.1	297.0^*, 285.0	35	18, 20
	37	wortmannin (渥曼青霉素)	8.36	[M+H]⁺	429.0	355.1^*, 313.1	18	12, 18

Group	No.	Compound	Retention time/min	Adduct	Precursor ion (m/z)	Product ions (m/z)	CV/V	CE/eV
*Quantification ion.
A	1	aflatoxin B₁ (黄曲霉毒素B₁)	8.25	[M+H]⁺	313.1	284.9^*, 241.0	46	26, 40
	2	aflatoxin B₂ (黄曲霉毒素B₂)	7.92	[M+H]⁺	315.2	287.1^*, 259.0	46	30, 32
	3	aflatoxin G₁ (黄曲霉毒素G₁)	7.73	[M+H]⁺	329.1	242.9^*, 200.0	42	28, 42
	4	aflatoxin G₂ (黄曲霉毒素G₂)	7.39	[M+H]⁺	331.1	189.0^*, 245.0	48	43, 34
	5	sterigmatocystin (杂色曲霉素)	10.81	[M+H]⁺	324.8	310.0^*, 280.8	45	26, 46
B	6	ochratoxin A (赭曲霉毒素A)	10.61	[M+H]⁺	404.1	238.9^*, 220.9	32	28, 40
	7	ochratoxin B (赭曲霉毒素B)	9.83	[M+H]⁺	370.2	204.9^*, 102.8	25	25, 54
	8	T-2 toxin (T-2毒素)	9.63	[M+NH₄]⁺	484.2	185.1^*, 305.3	24	25, 18
	9	T-2 triol (T-2三醇)	8.34	[M+Na]⁺	405.2	303.1^*, 125.0	62	14, 16
	10	HT-2 toxin (HT-2毒素)	9.03	[M+NH₄]⁺	442.2	263.2^*, 245.1	30	10, 10
	11	lysergol (麦角醇)	4.32	[M+H]+	254.8	197.0^*, 44.0	15	24, 15
	12	ergocornine (麦角柯宁碱)	7.72	[M+H]⁺	562.2	208.0^*, 223.1	36	48, 40
	13	zearalenone (玉米赤霉烯酮)	10.83	[M-H]^-	317.2	131.0^*, 175.1	48	35, 30
	14	α-zearalenol (α-玉米赤霉烯醇)	10.96	[M-H]^-	319.0	275.1^*, 301.1	44	33, 30
	15	β-zearalenol (β-玉米赤霉烯醇)	10.34	[M-H]^-	319.0	275.1^*, 301.1	42	32, 28
	16	zearalanone (玉米赤霉酮)	10.70	[M-H]^-	319.0	275.1^*, 301.1	43	28, 34
	17	α-zearalanol (α-玉米赤霉醇)	10.69	[M-H]^-	321.0	277.2^*, 303.2	42	26, 26
	18	β-zearalanol (β-玉米赤霉醇)	10.11	[M-H]^-	321.0	277.2^*, 303.2	40	28, 32
C	19	deoxynivalenol (脱氧雪腐镰刀菌烯醇)	4.38	[M+H]⁺	296.9	249.1^*, 231.1	15	10, 15
	20	15-acetyldeoxynivalenol	6.75	[M+H]⁺	338.6	137.0^*, 321.0	25	22, 15
		(15-乙酰基脱氧雪腐镰刀菌烯醇)
	21	3-acetyldeoxynivalenol	6.75	[M+H]⁺	339.0	231.0^*, 203.1	28	16, 22
		(3-乙酰基脱氧雪腐镰刀菌烯醇)
	22	fumonisin B₁ (伏马毒素B₁)	8.69	[M+H]⁺	722.2	352.3^*, 334.3	50	36, 40
	23	fumonisin B₂ (伏马毒素B₂)	9.49	[M+H]⁺	706.4	336.5^*, 354.5	40	40, 35
	24	fumonisin B₃ (伏马毒素B₃)	9.14	[M+H]⁺	706.4	336.5^*, 354.5	40	40, 34
	25	fusarenon X (镰刀菌烯酮-X)	5.65	[M+H]⁺	355.1	247.1^*, 175.1	24	14, 22
	26	diacetoxyscirpenol (蛇形菌素)	8.07	[M+NH₄]⁺	384.2	307.2^*, 105.0	25	15, 30
	27	neosolaniol (新茄病镰刀菌烯醇)	6.09	[M+NH₄]⁺	400.2	185.0^*, 215.1	24	25, 20
	28	tentoxin (腾毒素)	9.32	[M+H]⁺	415.2	312.2^*, 256.1	66	20, 28
	29	patulin (展青霉素)	2.30	[M+H]⁺	155.0	71.0^*, 81.0	30	10, 10
	30	glotoxin (胶霉毒素)	8.30	[M+H]⁺	327.4	263.1^*, 245.1	26	14, 22
	31	verruculogen (疣孢青霉原)	10.71	[M+Na]⁺	534.0	392.1^*, 435.2	30	18, 20
	32	penitrem A (青霉震颤素A)	11.78	[M+H]⁺	634.3	558.3^*, 616.3	22	18, 10
	33	penicillic acid (青霉酸)	5.46	[M+H]⁺	171.0	97.0^*, 82.0	20	17, 25
	34	mycophenolic acid (霉酚酸)	9.69	[M+H]⁺	320.9	207.0^*, 159.0	25	26, 32
	35	roquefortine C (娄地青霉素C)	7.89	[M+H]⁺	389.9	193.0^*, 108.2	40	30, 42
	36	citreoviridin (黄绿青霉素)	10.38	[M+H]⁺	403.1	297.0^*, 285.0	35	18, 20
	37	wortmannin (渥曼青霉素)	8.36	[M+H]⁺	429.0	355.1^*, 313.1	18	12, 18

No.	Matrix effects							LOQ/ (μg/kg)	Range/ (μg/L)
No.	Corn	Wheat flour	Cotton meal	Soybean meal	Pig concentrated feed	Pig compound feed	Cattle concentrate supplement	LOQ/ (μg/kg)	Range/ (μg/L)
Nos. are consistent with those of mycotoxins inTable 1.
1	0.89	1.03	0.61	0.87	0.98	0.88	1.02	2	0.5-25
2	0.90	1.00	0.76	0.62	1.21	0.81	1.70	2	0.5-25
3	0.95	1.01	0.83	0.99	0.87	0.88	0.65	2	0.5-25
4	0.95	1.04	0.87	0.98	1.21	0.90	0.46	2	0.5-25
5	0.72	0.76	0.59	0.76	0.97	0.72	0.88	2	0.5-25
6	0.94	0.92	0.87	1.06	1.01	0.95	1.01	20	5-250
7	0.98	0.94	0.98	1.18	1.31	0.91	1.11	20	5-250
8	0.87	0.82	0.77	0.83	0.91	0.68	0.82	20	5-250
9	1.13	0.96	0.38	0.97	0.18	0.82	0.04	20	5-250
10	1.01	0.98	0.63	0.32	0.84	0.38	0.78	20	5-250
11	0.89	0.94	0.76	1.44	2.07	0.96	1.94	20	5-250
12	0.36	0.37	0.22	0.28	0.39	0.68	0.33	20	5-250
13	0.82	0.93	0.81	0.99	0.94	1.18	1.12	20	5-250
14	0.87	1.14	0.82	1.03	1.15	1.57	1.29	20	5-250
15	0.85	0.97	0.78	1.03	0.67	1.22	1.05	20	5-250
16	0.81	0.94	0.80	1.01	1.03	1.11	1.21	20	5-250
17	0.87	1.10	0.89	1.12	0.86	1.28	1.18	20	5-250
18	0.86	1.07	0.77	1.07	1.06	1.15	0.97	20	5-250
19	0.81	1.03	0.67	1.00	1.02	0.75	0.70	40	10-500
20	0.79	0.87	0.62	0.79	0.87	0.40	0.66	40	10-500
21	0.79	0.90	0.61	0.84	0.89	0.46	0.66	40	10-500
22	2.36	1.47	2.54	1.13	1.00	3.84	1.17	40	10-500
23	1.72	1.78	1.76	0.94	1.03	2.14	1.04	40	10-500
24	1.31	1.71	2.06	1.06	1.16	3.55	1.37	40	10-500
25	1.02	1.03	0.84	0.96	1.05	0.84	0.59	40	10-500
26	0.99	0.97	0.86	0.97	1.51	0.89	1.66	40	10-500
27	1.04	1.00	0.94	1.02	1.19	0.91	0.65	40	10-500
28	0.91	0.96	0.79	0.96	1.15	0.84	0.91	40	10-500
29	0.89	0.98	1.00	0.89	1.02	0.84	0.76	40	10-500
30	0.46	0.50	0.27	0.22	1.16	0.30	1.20	40	10-500
31	0.84	0.90	0.56	1.27	0.26	1.09	1.08	40	10-500
32	0.59	0.66	0.53	0.70	0.76	0.77	0.80	40	10-500
33	1.00	1.03	0.98	1.04	1.83	0.90	0.96	40	10-500
34	1.04	0.93	0.91	1.06	1.63	0.98	1.57	40	10-500
35	0.62	0.87	0.59	0.78	0.54	0.71	0.49	40	10-500
36	0.83	0.89	0.71	1.24	1.61	0.97	1.21	40	10-500
37	0.73	0.80	0.22	0.64	0.47	0.58	0.64	40	10-500

No.	Matrix effects							LOQ/ (μg/kg)	Range/ (μg/L)
No.	Corn	Wheat flour	Cotton meal	Soybean meal	Pig concentrated feed	Pig compound feed	Cattle concentrate supplement	LOQ/ (μg/kg)	Range/ (μg/L)
Nos. are consistent with those of mycotoxins inTable 1.
1	0.89	1.03	0.61	0.87	0.98	0.88	1.02	2	0.5-25
2	0.90	1.00	0.76	0.62	1.21	0.81	1.70	2	0.5-25
3	0.95	1.01	0.83	0.99	0.87	0.88	0.65	2	0.5-25
4	0.95	1.04	0.87	0.98	1.21	0.90	0.46	2	0.5-25
5	0.72	0.76	0.59	0.76	0.97	0.72	0.88	2	0.5-25
6	0.94	0.92	0.87	1.06	1.01	0.95	1.01	20	5-250
7	0.98	0.94	0.98	1.18	1.31	0.91	1.11	20	5-250
8	0.87	0.82	0.77	0.83	0.91	0.68	0.82	20	5-250
9	1.13	0.96	0.38	0.97	0.18	0.82	0.04	20	5-250
10	1.01	0.98	0.63	0.32	0.84	0.38	0.78	20	5-250
11	0.89	0.94	0.76	1.44	2.07	0.96	1.94	20	5-250
12	0.36	0.37	0.22	0.28	0.39	0.68	0.33	20	5-250
13	0.82	0.93	0.81	0.99	0.94	1.18	1.12	20	5-250
14	0.87	1.14	0.82	1.03	1.15	1.57	1.29	20	5-250
15	0.85	0.97	0.78	1.03	0.67	1.22	1.05	20	5-250
16	0.81	0.94	0.80	1.01	1.03	1.11	1.21	20	5-250
17	0.87	1.10	0.89	1.12	0.86	1.28	1.18	20	5-250
18	0.86	1.07	0.77	1.07	1.06	1.15	0.97	20	5-250
19	0.81	1.03	0.67	1.00	1.02	0.75	0.70	40	10-500
20	0.79	0.87	0.62	0.79	0.87	0.40	0.66	40	10-500
21	0.79	0.90	0.61	0.84	0.89	0.46	0.66	40	10-500
22	2.36	1.47	2.54	1.13	1.00	3.84	1.17	40	10-500
23	1.72	1.78	1.76	0.94	1.03	2.14	1.04	40	10-500
24	1.31	1.71	2.06	1.06	1.16	3.55	1.37	40	10-500
25	1.02	1.03	0.84	0.96	1.05	0.84	0.59	40	10-500
26	0.99	0.97	0.86	0.97	1.51	0.89	1.66	40	10-500
27	1.04	1.00	0.94	1.02	1.19	0.91	0.65	40	10-500
28	0.91	0.96	0.79	0.96	1.15	0.84	0.91	40	10-500
29	0.89	0.98	1.00	0.89	1.02	0.84	0.76	40	10-500
30	0.46	0.50	0.27	0.22	1.16	0.30	1.20	40	10-500
31	0.84	0.90	0.56	1.27	0.26	1.09	1.08	40	10-500
32	0.59	0.66	0.53	0.70	0.76	0.77	0.80	40	10-500
33	1.00	1.03	0.98	1.04	1.83	0.90	0.96	40	10-500
34	1.04	0.93	0.91	1.06	1.63	0.98	1.57	40	10-500
35	0.62	0.87	0.59	0.78	0.54	0.71	0.49	40	10-500
36	0.83	0.89	0.71	1.24	1.61	0.97	1.21	40	10-500
37	0.73	0.80	0.22	0.64	0.47	0.58	0.64	40	10-500

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