Bacillus monnieri, (L.) (L.)

Bhandari, Pamita, Sendri, Nitisha & Devidas, Shinde Bhagatsing, 2020, Dammarane triterpenoid glycosides in Bacopa monnieri: A review on chemical diversity and bioactivity, Phytochemistry (112276) 172, pp. 1-14 : 2-3

3.1. Chromatographic analysis of B. monnieri

Despite the isolation and characterization of specialized metabolites, dammarane triterpenoid glycosides were analysed in natural matrixes of the plant and different herbal formulations by using different analytical techniques ( Renukappa et al., 1999; Ganzera et al., 2004; Shrikumar et al., 2004; Deepak et al., 2005; Murthy et al., 2006; Agrawal et al., 2006; Bhandari et al., 2006a,b, 2009a; Zehl et al., 2007; Srivastava et al., 2012; Ahmed et al., 2015). The various techniques used for analysis of specialized products in B. monnieri include TLC, HPLC (high performance liquid chromatography), HPTLC (high performance thin layer chromatography), SFC (super critical fluid chromatography) and LC-MS/MS (liquid chromatography mass spectrometry). The spectrophotometeric methods had also been developed for the quantification of bacosides in which bacosides were hydrolysed and aglycone was measured at 278 nm ( Pal and Sarin, 1992). A HPLC-PDA method was described for the quantification of six bacosides and enlisted that the total saponin contents varied in the range of 1.06–13.03%. The findings showed that accumulation of dammarane triterpenoid glycosides was more in the leaves as compared to the stem part of the herb ( Ganzera et al., 2004). For the determination of bacoside-A3 and bacopaside –I content in B. monnieri extract and monoherbal formulations, a HPTLC and SFC methods were developed by different research groups ( Shrikumar et al., 2004; Agrawal et al., 2006). As discussed previously that the structures of bacoside A & B were conflicting and most of the pharmacological and clinical studies of the B. monnieri extract were considered due to bacoside A and bacoside B ( Chatterji et al., 1963; Chatterji et al., 1965; Kawai and Shibata, 1978). To resolve this ambiguity, authors attempted to establish the chemical structures of bacoside A and accomplished that it was a mixture of four saponins ( Deepak et al., 2005). Later on, the composition of bacoside B was also eastablished as a mixture of four diglycosidic saponins ( Sivaramakrishna et al., 2005b). An analytical HPLC–DAD method had been developed for quantification of twelve saponins in plant extract and herbal formulations in which saponins were resolved on Luna C18 column (Phenomenex) with isocratic elution of 0.05 M sodium sulphate buffer and acetonitrile (68.3:31.5, v/v) with flow rate of 1.0 ml/min ( Murthy et al., 2006). The results revealed that bacopaside II, bacopaside I, bacopaside X and bacopasaponin C dominate the bacopasaponin E and F. Bhandari et al., 2009 reported a high performance liquid chromatographic method using silica-based monolithic column coupled with evaporative light scattering detector (HPLC-ELSD) for quantification of bacosides (bacoside A, bacopaside I, bacoside A 3, bacopaside II, bacopaside X, bacopasaponin C) and apigenin in nine accessions of B. monnieri . The method was developed on five different reversed phase analytical columns and found that optimum separation of analytes was achieved on chromolith column. The same authors standardized a method for quantification of bacoside A in minimum amount of plant material i.e. up to 2 mg by HPLC ( Bhandari et al., 2006a,b). Afterwards number of HPTLC methods were developed for quantification of bacosides in B. monnier i ( Ahmed et al., 2015; Nuengchamnong et al., 2016).

Furthermore, for rapid screening of specialized metabolites in B. monnieri , the techniques like HPLC coupled with mass spectrometry (HPLC-MS), and nuclear magnetic resonance spectroscopy (LC-NMR) had also been used. For the qualitative analysis of dammarane triterpenoid glycosides, a descriptive reversed phase high performance liquid chromatography coupled with nuclear magnetic resonance and mass spectroscopy (LC-NMR-MS) was used on LiChrosphere RP-18 HPLC column using acetonitrile-water as mobile phase with a flow rate of 0.8 ml/min ( Renukappa et al., 1999). In the analysis, four major peaks were observed, of which three peaks were identified as bacoside A 3, 3- β -[O -β -D-glucopyranosyl (1 → 3)- O -[α -L-arabinofuranosyl-(1 → 2)]- O -β -D-arabinopyranosyl) oxy] jujubogenin and bacopasaponin C. In another study, different MS techniques like ESI-ion trap (IT), AP-MALDI-IT, MALDI-IT/reflectron time-of-flight (RTOF) MS, all utilizing low-energy collision-induced dissociation (CID) and high-energy CID analysis were used to understand the glycosidic linkages and distinction of aglycone ( Zehl et al., 2007). The fragmentation behaviour observed from these techniques indicated that all the applied techniques produced the sequence and branching of the glycan and the molecular mass of the aglycone. Although low-energy collision induced dissociation (CID) provides the information about the interglycosidic linkages however, this study was not able to differentiate the type of aglycone (jujubogenin or pseudojujubojenin). In another study, Srivastava et al. analysed the B. monnieri for bacopaside-I and bacoside A content and suggested that the maximum yield of bacosides can be achieved at temperature up to 60 °C. To study the effect of geographical variations, a HPTLC method was used to study the content of bacoside A and concluded that highest content of bacoside A was observed in leaves collected from Jammu region while lowest content was examined in sample collected from Kerela region. In Addition to this, a liquid chromatography coupled with electrospray ionization quadrupole time-of-flight mass spectrometery (LC-ESI-QTOF-MS) was conducted to characterize dammarane triterpenoid glycosides in B. monnieri ( Nuengchamnong et al., 2016) . In this study authors reported that the aglycone (jujubogenin and pseudojujubogenin) of B. monnieri can be distinguished on the fragmentation pattren.