Method:

HPLC

Matrix:

Healthy products

Application No.:

101919

Columns:

Cat No.:

87003

Conditions:

Column: EndeavorsilTM C18，100×2.1 mm，1.8 μm (Cat#:87003)
Mobile Phase A: 0.05 mol/L sodium acetate (adjusted to pH=6.50±0.05 with glacial acetic acid)
Mobile Phase B: Methanol : Acetonitrile : H2O=20 : 60 : 20(v/v/v)
Flow Rate: 0.3 mL/min
Detector: UV 254 nm
Column Temp*: 35 ℃
Injection Volume: 2.0μL
Sample Solution: Pipetted 200 μL mixed solution(19 Natural Amino Acids +NH3; concentration of Amino Acids is 2.0
mmol/L, excepted concentration of Cystine is 1.0 mmol/L) and derivatization solution(100 μL 1 mol/L triethylamine-ACN solution+100 μL 0.2 mol/L PITC-ACN solution) into a 1.5 mL centrifuge tube, mixed thoroughly and reacted for 1 h
at ambient temperature. Then added 400 μL n-hexane and shaked gently for 5-10 s, rested for a while, pipetted 200
μL lower solution and mixed with 800 μL H2O, filtered with 0.22 μm syringe filter.

Gradient Procedures
Time/min 0.0 5.6 5.8 8.0 8.5 12.0
A/% 95 52 0 0 95 95
B/% 5 48 100 100 5 5

* Must keep column temperature at 35 ℃.

Author:

Dikma Technologies Inc.

Table:

1 Aspartate 2 Glutamate 3 Serine 4 Glycine
5 Histidine 6 Arginine 7 Threonine 8 Alanine
9 Proline 10 Tyrosine 11 Valine 12 Methionine
13 Cystine 14 Isoleucine Ile 15 Leucine 16 Norleucine
17 Phenylalanine 18 Tryptophan 19 Lysine