Development of an LC-MS/MS-based analytical method for quantification of soybean allergen Gly m 4 in soybean grains and processed foods

Figures & data

Table 1. List of peptide sequences and ions used for the MRM transitions.

Peptide	Precursor (charge state), m/z	Product ions (fragment), m/z		Collision energy (V)
		1	2
ALVTDADNVIPK	628.67 (+2)	244.2 (y2)	284.3 (b3)	25

Figure 1. LC-MRM chromatograms of the ALVTDADNVIPK peptide of representative trypsin digested samples. Chromatograms show the selectivity for (a) 10 µg/mL of Gly m 4, (b) soybean grain, (c) roasted soybean flour and (d) soybean milk.

Figure 2. A representative calibration curve of trypsin-digested rGly m 4.

Table 2. Percent of recovery and the coefficients of variation.

Spiked rGly m 4 concentration (µg/mL)	Recovery (%)	Intra-assay CV (%)	Inter-assay CV (%)
10	128	13.2	12.8

Table 3. Specificity of the LC-MS/MS-based method.

Foods	Gly m 4 homologue concentration (µg/g)
Adzuki bean grain	N.D.^a)
Red kidney bean grain	N.D.^a)
Chickpea grain	N.D.^a)
Lentil grain	N.D.^a)
Corn grain	N.D.^a)
Mixed juice of fruits and vegetables	N.D.^b)

^a)N.D.: <0.5 μg/g in Gly m 4 standard curve. n = 1.

^b)N.D.: <0.1 μg/g in Gly m 4 standard curve. n = 1.

Table 4. Levels of Gly m 4 in soybean grains and soybean processed foods.

Foods	Manufacturer No.^a)	Gly m 4 concentration, mean ± SD (µg/g)^b)
Soybean	1	421 ± 4.4
	2	343 ± 13.6
	3	333 ± 27.8
	4	309 ± 6.6
	5	332 ± 10.4
Roasted soybean flour	1	45 ± 1.4
	2	2.7 ± 0.14
Soybean milk	1	60 ± 2.0
	2	67 ± 2.0
	3	45 ± 2.2

^a)Numbers indicate different manufacturers.

^b)Value was determined in n = 3.