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To establish a liquid chromatography-tandem mass spectrometry (LC-MS/MS) method for determining the concentration of cyclic guanosine monophosphate (cGMP) in lysates of human colon adenocarcinoma lung metastasis cells (T84 cells) after co-incubation with linaclotide.
A Kromasil 100-5-C18 column (150 mm×2.1 mm, 5 μm) was used with a mobile phase consisting of 0.1% formic acid in water and 0.1% formic acid in methanol, employing a gradient elution at a column temperature of 50 ℃. The detection was performed using an electrospray ionization (ESI-) source and multiple reaction monitoring (MRM) mode, with the monitored ion transitions for cGMP and the internal standard 8-Br-cGMP being m/z 344.20 → 150.00 and m/z 423.90 → 230.00, respectively.
The linear ranges for cGMP were 1 to 500 ng·mL-1 (r≥ 0.999). The method demonstrated precision, accuracy, matrix effects, and extraction recovery rates that met analytical requirements. The method was successfully applied to accurately detect cGMP levels in cells after administration of two types of linaclotide capsule formulations. A significant concentration-dependent change in cGMP levels was observed after co-incubation of linaclotide with T84 cells for 30 min, with EC50 values of 167.6 and 112.1 nmol·L-1 for the reference and test formulations, respectively.
The method established in this study demonstrates excellent selectivity and accuracy, effectively quantifying cGMP levels in lysates of human colon adenocarcinoma lung metastasis cells, providing a reliable analytical tool for related pharmacological research.
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| 科 Family | 属数 Number of genus | 种数 Number of species | 占总种数比例 Percentage of total species (%) | 属 Genus | 种数 Number of species | 占总种数比例 Percentage of total species (%) |
|---|---|---|---|---|---|---|
| 鹅膏菌科Amanitaceae | 2 | 11 | 5.26 | 鹅膏菌属 Amanita | 10 | 4.78 |
| 小菇科 Mycenaceae | 2 | 12 | 5.74 | 丝盖伞属 Inocybe | 5 | 2.39 |
| 多孔菌科 Polyporaceae | 8 | 14 | 6.70 | 蜡蘑属 Laccaria | 5 | 2.39 |
| 红菇科 Russulaceae | 3 | 23 | 11.00 | 小皮伞属 Marasmius | 6 | 2.87 |
| 小菇属 Mycena | 11 | 5.26 | ||||
| 光柄菇属 Pluteus | 5 | 2.39 | ||||
| 红菇属 Russula | 17 | 8.13 | ||||
| 栓菌属 Trametes | 5 | 2.39 |
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