Document Type

Thesis

Publication details

Casselman, I 2016, 'Genetics and phytochemistry of Salvia divinorum', PhD thesis, Southern Cross University, Lismore, NSW.

Copyright I Casselman 2016

Abstract

Salvia divinorum Epling & Játiva (S. divinorum) is endemic to the Sierra Mazateca cloud forests of Oaxaca, Mexico. It has been used as a traditional medicine and in ceremonies by the Mazatec, the indigenous people of Oaxaca, for many centuries. In more recent times Salvia divinorum has become globally recognized, both for its potent psychoactive effects, as well as its potential use as a phyto-medicine.

Previous research by the author demonstrated that S. divinorum use is a growing, global phenomenon, particularly for recreational use. Despite this, published, peer reviewed research on this plant species is limited in comparison to other medicinal plants. The purpose of this research was to explore the genetic and phytochemical variation of S. divinorum.

One study has examined intraspecifc variation of this species but only in a small number of Oaxacan samples. Since its global dispersal from Mexico, propagation has been primarily by clonal methods, therefore, genetic variation of this species outside Mexico may be limited. Before to commencing investigation of intraspecifc variation, it was necessary to assess the quality of DNA which could be extracted from fortifed Saliva divinorum material. It was confrmed that DNA of suffcient quality could be obtained from fortifed extracts for further intraspecifc analysis. Regions of the chloroplast genome shown to be variable within other plant species were amplifed with universal primers to assess genetic variation in S. divinorum. In particular, regions containing chloroplast simple sequence repeats (cpSSR's) were targeted as these have been widely used for the detection of intraspecifc variation and phylogeography in plants. Chloroplast sequence data, approximately 2.4 kilobases in length, including the intergenetic spacers trnS-trnG, rps16-trnK, trnL-trnF and the trnL intron, were examined for variation among S. divinorum samples from Europe, North America and Australia. Sequenced alignments contained cpSSR repeat regions, however, no genetic variation was detected in more than 40 globally-distributed plant samples and fortifed commercial products used in the study.

The majority of phytochemical research, to date, has focused on salvinorin A, the main psychoactive diterpene in S. divinorum, and little is known of other chemical compounds or chemical variation in S. divinorum. Numerous compounds have been identifed across the

Salvia genus and many of these may also be present but not identifed in S. divinorum. HPLC and LC/MS data generated from the collected samples was used to perform two principal component analyses (PCA) to elucidate patterning in the chemical make up of S. divinorum. This analysis revealed two similar sets of groupings in both PCAs. HPLC and LC/MS analysis confrmed the presence of rosmarinic acid in S. divinorum. This compound is commonly identifed in the Salvia genus and it is surprising that it has not been confrmed in S. divinorum previously, as the UV peak is very prominent on the chromatogram. A high variation in the concentration of rosmarinic acid was also observed across the 37 samples tested. Not all salvinorin A fortifed products had higher salvinorin A concentrations than unfortifed samples.

The variation in the phytochemical make up and the lack of genetic variation in S. divinorum are interesting results and the combined genetic phytochemical approach offers novel insights. The lack of genetic variation found, adds evidence to the proposition that all plants outside Mexico have a common source. While there are several other factors which may affect phytochemical variation it may be that chemical variation in S. divinorum is determined phenotypically. Growing conditions may be the primary consideration to achieve optimal levels of active constituents as this species is further developed as a phyto-medicine.

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