The objective was to determine how measurements of plasma free AA by gas chromatography (GC) after chloroformate derivatization compared with values obtained by ion-exchange chromatography (HPLC) followed by ninhydrin derivatization. Plasma was obtained from lactating dairy cows (mean DIM = 148) part of a trial were Met was abomasally infused (12 or 24 g Met/d) or fed in rumen-protected form (15 or 30 g Met/d). Blood samples (n = 89) were collected from coccygeal vessels into evacuated tubes. After centrifugation, plasma was split into 2 aliquots and stored frozen at −80°C. Each set was analyzed either by HPLC in a commercial laboratory, or in-house by GC using a kit (EZ:faast, Phenomenex). This method involved deproteinization with TCA, AA collection using solid phase extraction, derivatization with chloroformate, GC separation in a capillary column and flame ionization detection. Quantitation was based on area under the curve, using the internal standard ratio method (Norvaline). Data were analyzed by Pearson correlation (r; Table 1). However Bland-Altman posited that correlation is misleading to compare methods and instead proposed the inspection of differrence plots and limits of agreement. Calculation of concordance correlation coefficients provided a quantitative alternative (CCC; Table 1). Overall correlation between methods was 0.95 (P < 0.001); nevertheless, correlations varied greatly for individual AA from 0.97 for Met to 0.15 for Glu. For Met, most data were within the limits of agreement established by Bland-Altman approach. Furthermore among all AA, CCC for Met was the closest to 1 (indicating almost total agreement between methods; Table 1). For research focusing on Met, GC offers a cheaper and faster alternative to HPLC. Table 1. Free AA (except Arg and Trp) in plasma measured by two methods

Amino acid	GC (µM)	HPLC (µM)	r	CCC
His	50.2	62.8	0.42	0.18
Ile	107.6	101.6	0.96	0.94
Leu	121.2	145.2	0.92	0.36
Lys	72.4	80.8	0.35	0.67
Met	32.8	33.9	0.97	0.99
Phe	40.0	46.1	0.65	0.37
Thr	186.9	117.9	0.96	0.25
Val	348.0	266.1	0.89	0.25
Ala	351.7	267.9	0.88	0.21
Asn	44.0	57.1	0.84	0.26
Asp	9.4	3.9	0.51	0.07
Cys	18.9	21.0	0.35	0.56
Glu	56.1	39.7	0.15	0.08
Gln	110.6	200.6	0.78	0.10
Gly	463.3	348.8	0.92	0.36
Pro	102.2	81.4	0.84	0.25
Ser	127.1	95.3	0.89	0.32
Tyr	30.5	42.1	0.60	0.31