Simultaneous Quantification of Four Indole Alkaloids in Catharanthus roseus Cell Line C20hi by UPLC-MS

Abstract

An ultra-performance liquid chromatography/mass spectrometry method to simultaneously quantify vindoline, catharanthine, serpentine and ajmalicine in Catharanthus roseus cell line C20hi is reported. Samples were extracted with 1% acetic acid, basified to pH 10 with ammonia, then extracted with ethyl acetate, dried, reconstituted with methanol-1% acetic acid water solution (1:1, v/v) and analyzed using an acetonitrile-0.1% formic acid gradient as the mobile phase. Detection was carried out by electrospray ionization mass spectrometry in the positive-ion mode with selective ion monitoring. The analysis of one sample was achieved in 6 min. The limits of detection were 0.46 -0.70 ng/ml in cell samples, and 0.10 -0.16 ng/ml in medium samples. The linearity of detection was over the wide range of 1.00 -6250.0 ng/ml. Intra-and inter-day accuracies (recovery 88.0 -111.8%) and precision (RSD 1.25 -7.81%) showed the performance of the assay. This method provides a more sensitive and high-throughput technique to quantify the four alkaloids in large amount of samples, and will be helpful in high-production cultivar screening.

Enhancement of Vindoline Production in Suspension Culture of the Catharanthus roseus Cell Line C20hi by Light and Methyl Jasmonate Elicitation

“…Therefore, a more sensitive and rapid UPLC method was developed to quantify vindoline and catharanthine after elicitation, in which the limit of detection was 0.70 ng/ml for vindoline and 0.46 ng/ml for catharanthine. 18 As shown in Fig. 3A, the light treatment significantly increased vindoline production in C20hi cells by about 0.49 -5.51-fold compared to the controls (n = 3, p < 0.01, t-test).…”

“…Therefore, we developed a more sensitive and rapid method by UPLC-MS. As described, the limit of detection for vindoline by UPLC-MS was about 1000-times lower than that reported by traditional HPLC methods, and the total analysis time was reduced to 6.0 min. 18,24 Using the UPLC-MS method, we detected concentrations of as low as 1.63 ng/ml vindoline in unelicitated C20hi cells (Fig. 4), and found significantly increased vindoline production after elicitation with light or light combined with MJ ( Figs.…”

“…18 Before UPLC-MS quantification, the existence of vindoline and catharanthine was first confirmed using liquid chromatography/electrospray ionization tandem mass spectrometry (LC/ESI-MS n ). The LC/ESI-MS n analysis was performed to identify alkaloids in the m/z range of 250 -1000 on a Surveyor LCQ DECA XP plus system (Thermo Finnigan, San Jose, CA) equipped with an ESI interface and an ion-trap mass analyzer.…”

Topical treatment with Tong-Luo-San-Jie gel alleviates bone cancer pain in rats

“…The chemical profile of the formulated gel was established by ultra performance liquid chromatography (UPLC) analysis (He et al, 2011; Wang et al, 2011) employing gradient elution with acetonitrile and 0.1% formic acid as mobile phases A and B, respectively. The ESI source was operated in the positive ionization mode.…”

Analysis of Indole Alkaloids from Rhazya stricta Hairy Roots by Ultra-Performance Liquid Chromatography-Mass Spectrometry

“…However, the availability of authentic standard compounds is limited since only a few pure compounds present in R. stricta are commercially available and components must therefore be isolated and purified from crude extracts, which makes the identification complex and time consuming [4]. Recently, an improved chromatographic separation of alkaloids was achieved by the introduction of ultra-performance liquid chromatography (UPLC) [5,6,7]. In UPLC, the stationary phase particle size of less than 2.5 µm has been shown to result in a significant improvement in separation efficiency [8].…”