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Table 2 Optimal conditions for LC analysis of nucleosides

From: Rapid and selective simultaneous quantitative analysis of modified nucleosides using multi-column liquid chromatography-tandem mass spectrometry

LC system NANOSPACE SI-2 (SHISEIDO)
Analytical column CAPCELL PAK ADME
(100 × 2.1 mm i.d., 3-μm particle size, Shiseido)
Mobile phase A, 20 mM CH3COONH4/H2O
(Adjusted to pH 5.3 using CH3COOH)
B, 1.0 M CH3COONH4/MeCN/H2O/CH3COOH
=1:95:5:0.03 (v/v/v/v)
Analytical gradient 0.0–1.0 min; A/B = 100.0/0.0
1.0–4.0 min; A/B = 100.0/0.0 → 91.4/8.6
4.0–5.0 min; A/B = 91.4/8.6 → 88.6/11.4
5.0–8.0 min; A/B = 88.6/11.4 → 79.9/20.1
8.0–10.0 min; A/B = 79.9/20.1 → 0.0/100.0
Analytical flow rate 0.0–10.0 min; 400 μL/min
Wash gradient 0.0–4.0 min; A/B = 0.0/100.0
4.0–5.0 min; A/B = 0.0/100.0 → 100.0/0.0
5.0–10.0 min; A/B = 100.0/0.0
Wash flow rate 0.0–1.0 min; 400 μL/min → 700 μL/min
1.0–4.0 min: 700 μL/min
4.0–5.0 min; 700 μL/min → 400 μL/min
5.0–10.0 min; 400 μL/min
Post-column addition MeOH; 400 μL/min
MS divert valve 0.0–0.5 min; waste
0.5–10.0 min; detector
Oven temperature 40 °C
Injection volume 3 μL