40
For more information, or to download product instructions,
Thermo Scientific
High-Purity Pre-Column Derivatization Reagents
PITC (Phenylisothiocyanate)
Ideal for the quantitative pre-column derivatization of
amino acids by reverse-phase HPLC.
14
N
C
S
PITC
Edman‘s Reagent
MW 135.19
Thermo Scientific PITC, also known as Edman’s Reagent,
reacts readily with amino acids in 5-10 minutes at room
temperature. The resulting phenylthiocarbamyl
derivatives can be separated and quantified in 30 minutes
using reverse-phase HPLC. This method produces stable
products with all amino acids, including proline.
TO COUPLE AMINO ACID STANDARD H WITH PITC.
1
1. Dry 10 µl Amino Acid Standard H in a small test tube. Dissolve dried
standard in 100 µl coupling buffer (acetonitrile: pyridine: triethylamine:
H
2
0, 10:5:2:3).
2. Dry standard solution by rotary evaporation. Dissolve the residual amino
acids once more in 100 µl of coupling buffer.
3. Add 5 µl of PITC.
4. Allow a 5-minute reaction at room temperature.
5. Evaporate sample to dryness by rotary evaporation under high vacuum.
6. Dissolve the resulting PITC-amino acids in 250 µl of 0.05 M ammonium
acetate, water or water:acetonitrile (7:2).
7. Analyze quantities of 1 to 10 µl (100 to 1,000 picomoles of each amino acid)
by reverse-phase HPLC.
NOTE:
Make certain that all HCI is evaporated before derivatization.
References
1. Heinrikson, R.L. and Meridith, S.C. (1984).
Anal. Biochem.
136
, 65-74.
2. Scholze, H. (1985).
J. Chromatogr.
350
, 453-460.
3. Janssen,
et al.
(1986).
Chromatogr.
22(7-12)
.
4. Evert, R.F. (1986).
Anal. Biochem.
154
, 431-435.
Ordering Information
Product # Description
Pkg. Size
TS-26922
PITC (Phenylisothiocyanate)
10 x 1 ml ampules
TS-20088
Amino Acid Standard H
10 x 1 ml ampules
