Initial TLC study of an uncharacterised Sceletium ('Little Karoo') Increasing societal and medicinal demand for S. tortuosum is seen in the West as both a widespread 'mood elevator', and gradually, more clinically, as an antidepressant option as it offers a novel dual action pharmacological profile of serotonin reuptake inhibition (other sources claim VMAT-2 mediated monoamine releasing properties) and PDE4 inhibition. It has been used both culturally in Africa and more recently spread worldwide without significant concerns with regard to either abuse potential or toxicity. Sceletium sp. Little Karoo is an undescribed species from the Little Karoo region in South Africa [1]. To date, little is known about it's potential phytochemistry, nor how it compares to more common Sceletiums such as Sceletium tortuosum or emarcidum which are culturally and medicinally important species, both used to prepare a 'Kanna' or Kougued product. Sceletium sp. Little Karoo as pictured by Herbalistics The plant grown from seed 1 There has been mention of many species of Sceletium including Sceletium strictum , Sceletium subvelutum (=Sceletium varians ), Sceletium tortuosum , Sceletium joubertii, Sceletium namaquense and others [2] The phytochemistry of various Sceletiums has been studied by Reddy and Stander et al. (2022) [3] to give the following profiles of alkaloidal constituents: Alkaloid constituents in various classes of Sceletium as analysed by Reddy, Stander et al. (2022) [3] Previous papers have studied more well known Sceletiums including S. tortuosum and S. emarcidum. The primary alkaloidal phytochemistry of Sceletium tortuosum is well characteristed, and centres around mesembrine-type alkaloids including mesembrine, mesembrenone and lesser alkaloids but individual S. tortuosum samples may vary considerably. The emarcidum -type Sceletiums are devoid of the parent mesembrine molecule but contain 4'-O-demethylated mesembrine analogues. 2 Various Sceletium alkaloids [4] A typical HPLC chromatogram of Sceletium tortuosum is shown below (Patnala and Kanfer, 2010) [5] showing characteristic peaks of constituents: 3 A totally characteristic variety of the Sceletiums is exemplified by S. varians which instead contains joubertiamine-type alkaloids, rather than mesembrine-type alkaloids. The pharmacology of these alkaloids is poorly studied. Structure of joubertiamine-type alkaloids Another distinct type of alkaloid found in Sceletium sp. is the tortuosamine-type alkaloids, similarly these are poorly studied with regard to CNS activity. 4 Structure of tortuosamine-type alkaloids In my work, I have subjected various Sceletium tortuosum and emarcidum samples to initial citizen TLC, the characteristic findings have been a constituent Rf ~0.3 which has been noted in diverse Mesembryanthemums known to be mesembrine containing. Other constituents have been noted in varous Sceletium tortuosum samples, including constituents Rf = 0.5 and a higher Rf constituent Rf = 0.8. Other analysis results have confirmed this pattern of constituents ranging from raw herb, to commercial extracts. Sceletium emarcidum showed a slightly different profile of constituents, Rf = 0.14. 0.28, 0.44 Sceletium tortuosum 5:1 extract (REF) and Sceletium emarcidum characteristic constituents via TLC (Silica, glass backed, 0.2mm, acetone elution, I 2 visualisation) The purpose of this citizen science was to compare S. tortuosum and S. emarcidum to 'Little Karoo' to see if it shared potential phytochemical similarity. Experimental Seeds were sourced from Herbalistics (Aus) and the plants grown locally. Collected plant material (leaf and stem material, collected winter) was dried at 70 deg. C and subjected to citizen TLC. The crushed, dried materials of all three Sceletiums (Little Karoo, S. tortuosum and S. emarcidum ) of approximately equal sample size were extracted with basified isopropanol (aq. ammonia) and concentrated to a small sample. Results: The Sceletium tortuosum sample, said to be 2.2% total alkaloids gave constituents of Rf = 5 0.13, 0.27, 0.33, 0.44 and a higher Rf band Sceletium emarcidum gave a slightly visibly band of Rf = 0.18 and a constituent at 0.4 Little Karoo gave a slightly pigmented band similarly to S. emarcidum at Rf = 0.18 and bands at Rf = 0.35 and 0.4 but comparatively weakly. TLC (Silica, glass backed, 0.2mm, acetone elution, I 2 visualisation). ST = S. tortuosum, SE = S. emarcidum and LK = Little Karoo. Discussion: The Little Karoo showed visibly pigmented low Rf similarities to the Sceletium emarcidum sample as well as constituents visible after I 2 visualisation which were fairly similar in the two species. That was in contrast to those seen in the S. tortuosum sample. That said, 6 visualisation was fairly faint in comparison to the S. tortuosum sample. There was some bands corresponding to the hypothesised mesembrine-type alkaloid Rf ~0.3 in the Little Karoo. Conclusion: Sceletium sp. Little Karoo may share some similarities to the emarcidum -type Sceletium and seems likely to contain active mesembrine-type alkaloids. References: [1] 'Sceletium sp. ‘Little Karoo’ – Kanna (seed)' herbalistics.com.au Retrieved June 2023 https://herbalistics.com.au/product/sceletium-sp-little-karoo-kanna-seed/ [2] Gericke, N., & Viljoen, A. M. (2008). Sceletium—A review update. Journal of Ethnopharmacology, 119(3), 653–663. https://doi.org/10.1016/j.jep.2008.07.043 [3] Reddy K, Stander M et al. Mass Spectrometry Metabolomics and Feature-Based Molecular Networking Reveals Population-Specific Chemistry in Some Species of the Sceletium Genus Front. Nutr., 29 March 2022, Food Chemistry, Volume 9 - 2022 | https://doi.org/10.3389/fnut.2022.819753 [4] Felix Makolo, Alvaro Viljoen, Clinton G.L. Veale, Mesembrine: The archetypal psycho ac Ɵ ve Scele Ɵ um alkaloid (2019) Phytochemistry, Volume 166, 2019, 112061 https://doi.org/10.1016/j.phytochem.2019.112061 [5] Patnala, S., & Kanfer, I. (2010). HPLC Analysis of Mesembrine-Type Alkaloids in Sceletium Plant Material Used as An African Traditional Medicine. Journal of Pharmacy & Pharmaceutical Sciences, 13(4), 558–570. https://doi.org/10.18433/J3DK5F 7