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Amino Acids (9-Fluorenylmethyl-Chloroformate Derivatives) on Raptor ARC-18

LC_FF0539
PeakstR (min)Conc.
(µmole/mL)
1.Arginine (Arg)5.1450.4
2.Asparagine (Asn)5.5180.4
3.Glutamine (Gln)5.6300.4
4.trans-4-Hydroxyproline (Hyp)5.8780.4
5.Serine (Ser)6.1390.4
6.Aspartic acid (Asp)6.2560.4
7.Glutamic acid (Glu)6.6960.4
8.Threonine (Thr)7.0270.4
9.Glycine (Gly)7.4660.4
10.Alanine (Ala)8.2270.4
11.Proline (Pro)8.8030.4
PeakstR (min)Conc.
(µmole/mL)
12.Methionine (Met)9.3240.4
13.Valine (Val)9.5370.4
14.Norvaline9.6560.4
15.Tryptophan (Trp)9.9100.4
16.Phenylalanine (Phe)10.1200.4
17.Isoleucine (Ile)10.2140.4
18.Leucine (Leu)10.2850.4
19.Cystine10.7490.2
20.Histidine (His)12.0450.4
21.Lysine (Lys)12.1740.4
22.Cysteine (Cys)12.4630.4
23.Tyrosine (Tyr)12.8010.4
Amino acids are analyzed as 9-fluorenylmethyl-chloroformate derivatives.
ColumnRaptor ARC-18 (cat.# 9314A1E)
Dimensions:100 mm x 3 mm ID
Particle Size:2.7 µm
Temp.:30 °C
Sample
Diluent:0.1 N HCl
Conc.: 0.4 μmole/mL for each amino acid (0.2 μmole/mL for cystine)
Inj. Vol.:1 µL
Mobile Phase
A:0.1% Formic acid + 20 mM ammonium formate in water
B:0.1% Formic acid + 10 mM ammonium formate in 90:10 acetonitrile:water
Time (min)Flow (mL/min)%A%B
0.000.88020
6.250.86040
9.000.84060
10.000.84060
13.000.80100
13.010.88020
15.000.88020
DetectorUV/Vis @ 265, 4.8 nm
InstrumentWaters ACQUITY UPLC H-Class
NotesDerivatization reaction: 50 μL amino acid mix + 100 μL 0.2 N borate buffer (pH 10.0) + 50 μL 15 mM 9-fluorenylmethyl-chloroformate solution + 50 μL acetonitrile

The injection can be performed after 5 minutes of reaction time.