Quantitative Analysis of Bioactive Carbazole Alkaloids in Murraya koenigii

Abstract

Carbazole alkaloids induce apoptosis in HL-60 cells through activation of the caspase-9/caspase-3 pathway and they are targeted as potential anticancer agents. Thus, the naturally occurring carbazole alkaloids become important as precursors for lead optimization in drug development. A method based on ultra performance liquid chromatography coupled with photodiode-array detection was developed using reverse phase isocratic elution with 85:15 acetonitrile and ammonium acetate buffer (5 mM). Seven samples of Murrya koenigii (L.) Spreng. from north-central India (Uttar Pradesh) were analyzed. All three targeted analytes, koenimbidine (mk1), koenimbine (mk2) and mahanimbine (mk3), were well separated within 4.0 min with linearity of the calibration curves (r 2 > 0.999). The limits of detection and quantification of mk1, mk2 and mk3 were 0.7, 0.4, 0.04 µg/mL and 2.14, 1.21, 0.12 µg/mL, respectively. The natural abundance of mk1, mk2 and mk3 was 0.06-0.20, 0.04-0.69 and 0.13-0.42%, w/w, respectively, in the dried powdered leaves, whereas, the tissue specific distribution of carbazole alkaloids was observed in the order of predominance, mk1 leaf>root>fruit>stem, mk2 fruit>leaf >stem>root, and mk3 fruit>leaf>root>stem. The developed method was validated for limits of detection and quantification, repeatability, accuracy, precision and stability. This is the first report on the natural abundance of the major carbazole alkaloids in M. koenigii and the method developed can be used in HPLC/UPLC systems.

Quantitative Analysis of Bioactive Carbazole Alkaloids in Murraya koenigii (L.) from Six Different Climatic Zones of India Using UPLC/MS/MS and Their Principal Component Analysis

“…CAs showed characteristic UV/VIS bands due to the presence of carbazole alkaloid skeleton which was further confirmed by mass spectral analysis. The purities of compounds were greater than 98 % according to the HPLC analysis and these were all used later as standard compounds for our reference [17,18] . Structures of CAs analyzed are shown in Figure 2.…”

“…The purities of compounds were greater than 98 % according to the HPLC analysis and these were all used later as standard compounds for our reference. [17,18] Structures of CAs analyzed are shown in Figure 2.…”

“…Out of which, compounds (mk279A, mk309, mk323, mk293A, mk347, mk331A, and mk331B) were earlier identified and isolated from M. koenigii by us. [17,18] Additional data for the previously non-reported compounds (mk263, mk279B, mk293B, and mk616) is shared in Supplementary Fig-ure S1 and δ values of 1 H and 13 C-NMR are given in Supplementary Table 1. LC/MS grade acetonitrile, methanol, and ammonium acetate were procured from Sigma Aldrich while ethanol was purchased from Merck Specialties Pvt.…”

“…[16] In our studies on this plant over five years, we have analyzed the natural abundance and tissue-specific distribution of some major CAs detected in the samples of MK plant from central India and found the CAs to be present in nearly all plant parts including the renewable parts like leaf. [17] Also, very recently we have published the bioactivities of methanolic leaf extract along with seven isolated major CAs from this plant and established the structure-activity relationship. [18] In the present study, we aimed to understand the differences in the content of CAs within the MK population from different geographical areas of all the six climatic zones of India.…”

“…In another study, the genetic diversity between wild samples of this plant from different geographical regions of India was explored, [14,15] and the putative gene families involved in carbazole alkaloids biosynthesis were identified [16] . In our studies on this plant over five years, we have analyzed the natural abundance and tissue‐specific distribution of some major CAs detected in the samples of MK plant from central India and found the CAs to be present in nearly all plant parts including the renewable parts like leaf [17] . Also, very recently we have published the bioactivities of methanolic leaf extract along with seven isolated major CAs from this plant and established the structure‐activity relationship [18] .…”