Library

Method Abstract 380.1

MA 380.1 / 30-MINUTE AMINO ACID ANALYSIS OF HYDROLYZED SAMPLES

Pickering Laboratories specializes in the manufacturing of cation-exchange columns and eluants for amino acid analysis. No other technique has been shown to match the reproducibility and selectivity of ion-exchange analysis with post-column Ninhydrin detection. Nor is there a chromatography technique that provides as much information; the 570/440 nm signal ratio for each amino acid is a constant and so offers information about peak purity.

Pinnacle PCX post-column derivatization system provides a unique opportunity to combine eluant gradient capabilities of modern HPLC instruments with column temperature gradients. As might be expected, this capability also reduces the analysis time.

We introduce an accelerated Amino Acid analysis method for hydrolyzed samples that utilizes temperature and eluant gradients.

METHOD

Analytical Conditions

Column: High-efficiency Sodium cation-exchange column, 4.6 x 110 mm, Catalog number 1154110T
Flow Rate: 0.6 mL/min
Mobile Phase: See method

Post-column Conditions

Post-column System: Pinnacle PCX
Reactor Volume: 0.5 mL
Temperature: 130 ˚C
Reagent: Trione Ninhydrin reagent
Flow Rate: 0.3 mL/min
Detection: UV/VIS 570 nm for primary amino acids and
     440 nm for secondary amino acids
 

METHOD FOR OXIDIZED FEEDS HYDROLYSATE SAMPLES

TIME

Na270 %

Na425 %

Na640 %

RG011 %

0

100

0

0

4.0

100

0

0

15.0

0

100 

0

0

16.0

0

100

0

31.0

0

100

0

31.1 0 0 0 100
33.0 0 0 0 100
33.1 100 0 0 0
40 100 0 0 0
         

COLUMN OVEN PROGRAM

TIME

TEMP ºC

0

55

12

55

17

70

32

70

33

55

 

 

 

chart-m-380-1
1 Aspartic Acid
2 Threonine
3 Serine
4 Glutamic Acid
5 Proline
6 Glycine
7 Alanine
8 Cystine
9 Valine
10 Methionine
11 Isoleucine
12 Leucine
13 Norleucine
14 Tyrosine
15 Phenylalanine
16 Histidine
17 Lysine
18 Tryptophan
19 Ammonia
20 Arginine
21 Cysteic Acid
22 Methionine
Sulfone

 

 

 

 

 

 

METHOD FOR PROTEIN HYDROLYSATE SAMPLES

TIME

Na315 %

Na425 %

Na640 %

RG011 %

0

100

0

0

4.0

100

0

0

15.0

0

100 

0

0

16.0

0

100

0

31.0

0

100

0

31.1 0 0 0 100
33.0 0 0 0 100
33.1 100 0 0 0
40 100 0 0 0
         

COLUMN OVEN PROGRAM

TIME

TEMP ºC

0

46

4

46

9

70

32

70

33

46

 

 

 

 chart-m-380-1b
1 Aspartic Acid
2 Threonine
3 Serine
4 Glutamic Acid
5 Proline
6 Glycine
7 Alanine
8 Cystine
9 Valine
10 Methionine
11 Isoleucine
12 Leucine
13 Norleucine
14 Tyrosine
15 Phenylalanine
16 Histidine
17 Lysine
18 Tryptophan
19 Ammonia
20 Arginine
21 Cysteic Acid
22 Methionine
Sulfone