Grace Catalog 2008

hplc applications
Proteins & Peptides/Reversed-Phase
1
4 5 8
7
2 3
6
PTH Amino Acids
PTH Amino Acids
9
10
0 2 4 6 8 10 12 14 Min.
11
12
13
1416
17
15
1. His
2. Cys Acid
3. Arg
4. Asp
5. Ser
6. Gln
7. Thr
8. Gly
9. Cys
10. Ala
11. Tyr
12. Met
13. Pro
14. Trp
15. Phe
16. Ile
17. Leu
Column: Vydac ® Denali ® , 5µm, 150 x 4.6mm (Part No. 238DE5415)
Mobile Phase: A: Acetonitrile:25mM Potassium Phosphate, pH3.3 (20:80)
B: Acetonitrile:25mM Potassium Phosphate, pH3.3 (80:20)
Flow Rate: 1.0mL/min
Detector: UV at 254nm
life science
Intensity (cps)
Column: DENALI 238DE5415 (5 µm, 4.6 mm i.d. x 150 mm)
Flow:
1.0 mL/min
Eluent: A = 20:80 ACN:25 mM potassium phosphate, pH 3.3
B = 80:20 ACN:25 mM potassium phosphate, pH 3.3
0 to 75% B over T17 15 min
Ambient (23 °C) 26.4
Absorbance, 254 nm T18-20
31.5 T26
5 µL of PTH amino acid standard T13 39.4 mixture
32.8
T5
1) His, 2) Cys acid, 3) Arg, 4) Asp, 5) Ser, 6) Gln, 54.4
7) Thr, 8) Gly, 9) Cys, 10) Ala, 11) Tyr, 12) Met,
T8
13) Pro, 17.414) Trp, 15) Phe, 16) Ile, 17) Leu
29.6
33.2
7.4
T20 T11
19.3 25.6
33.7
6.0e4 Gradient:
5.5e4 Temp:
5.0e4 Detection:
4.5e4 Sample:
4.0e4 Peaks:
3.5e4
3.0e4
2.5e4
2.0e4
1.5e4
1.0e4
5000.0
T7
7.7 16.6
TIC of Tryptic Digest of Green Fluorescent Protein (GFP): 150pmol Load on 1mm i.d.
22.8
38.7
40.1
5 10 15 20 25 30 35 40 45 50 55 60 65 70 Min.
T9
55.8
57.2 T29
60.2 64.7
Column:
Vydac ® , 250 x 1mm (Part No. 238EV51)
Mobile Phase: A: Acetonitrile:0.05% Trifluoroacetic Acid (2:98)
B: Acetonitrile:0.05% Trifluoroacetic Acid (90:10)
Gradient: Time: 0 60
%B: 5 65
Flow Rate: 50μL/min
Detector: ms: ABI QSTAR
Injection Volume: 25μL GFP digest
Mass Spectrum of Tryptic Digest of GFP
Intensity (counts)
6.5
6.0
5.5
5.0
4.5
4.0
3.5
3.0
2.5
2.0
1.5
1.0
T10
790.361
T5
T26
1220.139
1492.071
T8
826.410
T18-20
1012.464
T24
507.293
T17
T9
T13
T11
1050.523
1208.645
804.370
T7
655.384
T20
474.329 982.501
827.416
T19
523.221 1041.482 T4
579.315 1119.553
847.384
1312.674
429.090
752.342 1511.078
0.5
0.0
400 500 600 700 800 900 1000 1100 1200 1300 1400 1500 1600
m/z
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