ACE UHPLC and HPLC Columns
Amino Acid Enantiomer Separation of Seawater Samples
This method enables the quantification of free, dissolved combined, particulate and total amino acid enantiomers from seawater.
After hydrolysis, hydrolysates are evaporated, dissolved in borate buffer (pH 10) and centrifuged to remove flocculates. Samples
are derivatised with OPA/IBDC (N-isobutyryL -D-cysteine) and SMC (S-methyL -L -cysteine) added as internal standard. Enantiomer
elution order can be reversed by using IBLC (N-isobutyryL -L -cysteine)
45
30
15
0
L-Arg B-Ala
Standard sample
D-Ala = 34 nM
SMC
L-Val
D-Leu L-Ileu
0 10 20 30 40 50 60
Fluorescence detection (LU)
Time (min)
D-Asp L-Asp
D-Glu L-Glu = 178 nM
D-Ser L-Ser
Gly = 538 nM
L-Thr
L-His
GABA L-Ala
L-Tyr
D-Val
L-Met
L-Phe
L-Leu
L-Lys
45
30
15
0
Seawater sample
Seawater sample
SMC
L-Met
L-Phe
L-Leu
0 10 20 30 40 50 60
Fluorescence detection (LU)
Time (min)
D-Asx L-Asx
D-Glx L-Glx
D-Ser L-Ser
Gly
L-Thr
B-LA-Alarg
L-His
D-Ala
GABA L-Ala
L-Tyr
D-Val
L-Val
D-Leu L-Ileu
L-Lys
Application #AN3880
Conditions
Column: ACE UltraCore 5 SuperC18
Dimensions: 250 x 3.0 mm
Part Number: CORE-5A-2503U
Mobile Phase: A: 95% 40 mM KH2PO4 pH 6.15 in H2O + MeOH/MeCN (93:7 v/v)
B: 62% MeOH/MeCN (93:7 v/v) + 38% A
Gradient: Time (mins) %B
0.0 0
13.0 27
33.0 36
38.0 58
54.0 92
55.0 100
57.5 0
60.0 0
Flow Rate: 0.7 mL/min
Temperature: 45 °C
Detection: Fluorescence, λex 330 nm λem 450 nm
D-Asx = D-Asp + D-Asn
L -Asx = L -Asp + L -Asn
D-Glx = D-Glu + D-Gln
L -Glx = L -Glu + L -Gln
Time - Minutes
Reproduced with permission of Department of Chemistry and Biochemistry, Université de Moncton, New Brunswick, Canada
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