Table 2. Method Validation Results^a.

folate vitamer (detectors used for the quantification)	linearity and sensitivity			intraday variability (RSD %, n = 3)		interday variability (RSD %, n = 3 × 3)		recovery (%, mean ± standard deviation,n = 3)
	R2	LOD (ng/mL)	LOQ (ng/mL)	retention time	peak area	retention time	peak area	extraction buffer	whole-grain wheat flour	faba bean flour
10-HCO-H2folate (FL 360/460 nm)	0.964	7.3	24.3	0.21	3.28	1.52	10.66	85 ± 12	67 ± 10	49 ± 5
10-HCO-PGA (FL 360/460 nm)	0.991	5.5	18.3	0.27	6.16	0.57	9.06	118 ± 3	88 ± 3	87 ± 18
5-HCO-H4folate (UV 290 nm)	0.982	13.1	43.6	0.34	1.29	0.84	5.94	122 ± 12	102 ± 4	92 ± 7
5,10-CH+-H4folate (UV 360 nm)	0.998	7.2	23.9	0.11	3.49	0.92	8.62	119 ± 2	160 ± 7	163 ± 23
5-CH3-H4folate (FL 290/356 nm)	0.993	0.4	1.4	0.51	0.96	0.91	6.31	103 ± 2	81 ± 2	92 ± 4
H4folate (FL 290/356 nm)	0.986	0.7	2.3	0.36	1.10	0.92	3.65	86 ± 8	98 ± 3	83 ± 6
PGA (UV 290 nm)	0.974	6.4	21.4	4.77	2.14	3.64	10.17	95 ± 11	92 ± 6	83 ± 6
total folate								108 ± 4	94 ± 4	86 ± 6

^aA linear range of 80–1440 pg/injection (the injection volume was 10–30 μL) was chosen for each vitamer, except for 5-HCO-H₄folate, 5,10-CH⁺-H₄folate, and 10-HCO-H₂folate (320–1440, 160–1440 and 160–2880 pg/injection, respectively). The injection volume used to determine LOD (the limit of detection) and LOQ (limit of quantification) was 20 μL. RSD, relative standard deviation. The recovery test was carried out by spiking different matrices with 140 ng of each vitamer (280 ng for 10-HCO-H₂folate), and recovery (%) was calculated by the following equation: 100 × (the folate amount of the spiked matrices—the folate amount of the unspiked matrices)/the amount of folate used for spiking.