Abstract:

In this work, certification of L-citrulline in seeding watermelon extract by liquid chromatography-tandem
mass spectrometry (LC-MS-MS), and rapid determination of underivatized L-citrulline in seeding watermelon by high
performance liquid chromatography (HPLC) were established. Single factor and orthogonal experiments were used to
optimize the extraction conditions. LC-MS-MS analysis was carried out using Acuity C18 (50 mm × 2.1 mm, 1.7 μm) as the
stationary phase and a mixture of 10% acetonitrile and 90% methanol-water (50:50, V/V) as the mobile phase at a flow rate
of 0.2 mL/min with a column temperature of 40 ℃. The electrospray ionization (ESI) source was applied and operated in
a positive ion mode. The diagnostic product ions of L-citrulline were m/z 176.09/158.9 and m/z 176.09/112.9. Moreover,
the ion pair of m/z 176.09/158.9 was used for quantification. HPLC separation was performed suing Platisil ODS C18
(250 mm × 4.6 mm, 5 μm) as the stationary phase and 0.03 mmol/L phosphoric acid as the mobile phase at a flow rate of
0.7 mL/min with a column temperature of 30 ℃, and the detection wavelength was set at 202 nm. The results showed that
a good linear relationship was obtained for L-citrulline in the concentration range of 0.5–100 μg/mL with R2 = 0.999 9.
Average recoveries varied from 95.12% to 104.21% with RSD of 1.86%–4.75% (n = 3). The content of L-citrulline in
seeding watermelon was determined to be 0.656–2.563 mg/g.

Key words: seeding watermelon, L-citrulline, non-derivatization, liquid chromatography coupled to tandem mass spectrometry (LC-MS-MS), high performance liquid chromatography (HPLC)