Identification and quantification using HPLC-ES/MS of four pentacyclic oxindole alkaloids in Uncaria tomentosa wild type

Abstract

<i><b>Uncaria tomentosa</b></i> of the <i>Rubiaceae</i> family is a liana or woody vine that is found in Central and South America, which derives its nickname “<b>Uña de Gato</b>” or translated “<b>Cats Claw</b>” from its claw shaped thorns that resemble the claws of a cat. These thorns are used for the plant vine to climb and stretch to a height of 30 m. <i>U. tomentosa</i> naturally produces a range of secondary metabolites such as alkaloids, triterpenes, flavonoids, glycosides, and procyanidins. The metabolites in <i>U. tomentosa</i> are valuable in the treatment a number of diseases like bursitis, which is the inflammation of bursae (small sacs) of synovial fluid in the body, genital herpes, cancer, allergies, and those people infected with the HIV virus (4, 5). In the current research study we examine the concentrations of metabolites in the roots, leaves, and stems of <i>U. tomentosa</i> by establishing a method to accurately quantitate the concentrations of metabolites. In particular, we are interested in identifying and quantifying four principal pharmacologically active pentacyclic oxindole alkaloids: <i>Mitraphylline, Isomitraphylline, Uncaria C (pteropodine)</i>, and <i>Uncaria E (isopteropodine</i>. In order to optimize growth conditions, media with the essential composition of nutrients were established for growth of the wild type. <i>U. tomentosa</i> wild type plants were grown in different media including Murashige and Skoog (MSO), Vi Murashige and Skoog (!4 MS), Driver and Kuniyuki Walnut (DKW), Woody Plant Medium (WPM), Gamborg B5 medium (B5), Nitsch & Nitsch (N/N), Nitsch & Nitsch Charcoal, Schenk and Hilderbrant (SH), WHITE’S, and Homeostatic Soil (HSO) to observe optimal growth conditions. The plants were harvested and sent for side by side analysis of pentacyclic oxindole alkaloid (POA) content. The coupling of High Performance Liquid Chromatography (HPLC) and Electrospray Ionization Mass Spectrometry (ESI/MS) was the analytical method of choice because of its selectivity and sensitivity toward these POA’s (12, 13). We were able to compare HPLC-ESI/MS quantitative data on the cultivating properties of each medium used for micropropagation of <i>U. tomentosa</i> and found that the Vi MS was proficient enough to produce efficient plant growth and alkaloid production. Side by side comparison of different growth media has never been reported, thus this work may improve other cultivating procedures in the case of optimizing raw material for further research.