Rapid and quantitative determination of solanesol in Nicotiana tabacum by liquid chromatography–tandem mass spectrometry

Abstract

A rapid and sensitive liquid chromatography–tandem mass spectrometry (LC–MS/MS) method with multiple reaction monitoring (MRM) was developed for the determination of solanesol in Nicotiana tabacum. Sample preparation was performed by ultrasonic extraction with methanol for 20 min and then supernatant was extracted with hexane. The method used atmospheric pressure chemical ionization (APCI) detection in positive-ion mode. The separation of solanesol was performed on a Symmetry Shield™ RP18 column with a mixture of acetonitrile and isopropanol (1:1, v/v) containing 2 mM ammonium acetate as mobile phase. Quantification of solanesol was performed by the standard addition method. The limit of quantification (LOQ) and limit of detection (LOD) of solanesol were, respectively, 5.0 ng/ml (S/N = 10) and 1.5 ng/ml (S/N = 3). The relative standard deviations of peak area were 0.89 and 1.12% for intra-day and inter-day, respectively. The recoveries of solanesol ranged from 97.72 to 99.67% and the corresponding R.S.D.s were less than 2.7%. Analysis took 5 min, making the method suitable for rapid determination of solanesol in N. tabacum. The proposed method has been successfully applied to the analysis of solanesol in various organs of N. tabacum.

Introduction

Nicotiana tabacum belongs to the Solanaceae family and the plant is considered to be a good source of a large number of bioactive substances. Recently, the chemical compositions of N. tabacum have attracted considerable attentions in the world [1], [2], [3], [4]. Solanesol, a 45-carbon, all-trans-nonaprenol (see Fig. 1), was first isolated from flue-cured tobacco [5]. Solanesol itself can be used as antiulcer and hypertension treating agent [6], [7]. In addition, solanesol is a necessary medical intermediate in the industrial synthesis of coenzyme Q₁₀ [8], [9], [10], which is an excellent medicine in cardiovascular disease, cancer, atherosclerosis and so on [11], [12], [13], [14], [15].

Solanesol is in fact found in many plants from the Solanaceae family, one member of which is the Nicotiana genus. Other members of the family known to contain solanesol include tomato plants, potato plants, eggplants and pepper plants [16]. However, it was reported that the content of solanesol in N. tabacum was considerably higher than that in other plants and thus this plant represented the most convenient source for large-scale isolation of solanesol [17], [18], [19]. So it is very important to determine the content of solanesol in N. tabacum.

Many analytical methods including of column chromatography weight, thin-layer chromatography (TLC), coulomb's analysis, gas chromatography (GC) and high performance liquid chromatography (HPLC) have been documented for the determination of solanesol [20], [21], [22], [23], [24], [25], [26], [27], [28]. All of the methods above suffered from some limitations, such as, column chromatography weight method had low recovery, the precision obtained by TLC was poor, coulomb's analysis had significant error, the GC method was complicated by the interference of solanesenes, produced from the pyrolysis of solanesol at high temperatures in the GC oven and the breakdown of solanesol hindered the direct quantification of solanesol. The HPLC methods above had poor selective and sensitivity compared with the LC–MS/MS methods proposed in the study.

At present, liquid chromatography–tandem mass spectrometry (LC–MS/MS) has been accepted by more and more people as a useful method for identification and determination of compounds [29], [30]. Especially, it is very effective in the analysis of compounds from complex samples because of its low detection limit, high sensitivity and the possibility for short run time [31], [32], [33]. Signal suppression or enhancement of the target extracts by matrix components is a common phenomenon in LC–MS/MS analysis and should be considered [34]. Moreover, interfering matrix components can affect accuracy of the proposed method and maybe lead to some compromising or erroneous results [35]. In order to avoid the problems related with matrix effect, some authors preferred to select the optimization of sample preparation, the optimization of the chromatographic system and MS/MS detection [36], [37], [38]. In addition, some authors refered to the necessity of implementation of the standard addition method as a form of eliminating matrix effects [39], [40].

Solanesol is a nonaprenol containing a linear hydrocarbon chain, which is made up of nine isoprenoids. CoQ₁₀ is an ubiquinone, whose side chain is made up of 10 isoprenoids. Structurally, solanesol and CoQ₁₀ have some similarities. LC–MS/MS method with MRM have been reported for the determination of CoQ₁₀ in N. tabacum [41]. As far as we are aware, no report on determination of solanesol in N. tabacum by LC–MS/MS method up to now. Considering it, the aim of this study is to propose a validated LC–MS/MS method with multiple reaction monitoring (MRM) for separation and determination of solanesol in N. tabacum. Regarding that matrix components in N. tabacum are complex, in order to avoid the problems related with matrix effect, each step mentioned above (sample preparation, chromatographic system and MS/MS detection) was carefully optimized in the study. At the same time, the standard addition method was used as a quantification method to further minimize the matrix effect in the study. We have found this technique to be suitable for the rapid and sensitive quantification of solanesol in N. tabacum. Based on this work, the contents of solanesol in various organs of N. tabacum are determined and compared in this paper.