Alchemica Posted March 23, 2023 (edited) I feel the succulent dimension holds an interesting element in potentially having more therapeutically useful "in the real world" dimensions than more commonly touted plant medicines for their often more gentle but occasionally empathogenic edge. Also, the combination of often pro-cognitive (PDE4 inhibition seen with mesembrine-type alkaloids) with potent antidepressant effects meets an unmet demand. Part 1: Background and 'citizen analysis' using readily available materials Some Aizoaceae have been analysed [1]: I wanted to see if it was feasible to get some rough idea of the phytochemistry of a plant using only readily available materials eg. making it suitable for citizen science. I've been curious about simple TLC and paper chromatography as tools, particularly how feasible 'citizen TLC analysis' of plants is using a suitable paper, readily available solvent systems and simple visualisation techniques, compared to more conventional TLC. Can simple maceration of a medicinal plant in a readily available benignish solvent, concentration, spotting on common craft materials, developing the plate and OTC visualisation give some rough idea of the phytochemistry of a medicinal plant? Sceletium has in the past been analysed via conventional TLC [2, 3] and more advanced techniques [4] It took about 30 trials to find a readily available solvent system and semi-suitable paper - after many failed attempts, I found an 'etch art paper' that seemed the best I could find. I'm still to follow up with confirmation on silica TLC plates but it seems to give *a very rough idea* Sceletium tortuosum (unfermented) was used as a reference. This gives a main compound Rf = 0.5-0.54 on the craft paper. I particularly wanted to research D. bosseranum and Trichodiadema stellatum Method: Sample preparation: Samples were dehydrated at 70 deg. C and powdered. The samples were macerated in isopropanol. If no phenolic constituents were expected, NaOH q.s. was used to basify, otherwise basified with ammonia solution. The isopropanol solutions were concentrated to a small sample. Samples were spotted onto a plate, either a suitable craft paper, or silica gel plates. Acetone was used to elute the plate and I2 vapour (from OTC tincture) used as a visualisation technique. Crude technique - Craft Paper Note: this technique is limited in effectiveness and I suggest if you're that keen to play around with something like this, I'd suggest you just stick with silica gel plates to save A LOT of hassle. Delosperma bosseranum Sceletium tortuosum (unfermented) was compared to D. bosseranum. A compound with the same Rf as Sceletium was noted along with other constituents. Trichodiadema stellatum (syn. barbatum) Sceletium tortuosum (unfermented) was compared to D. bosseranum. A compound with the same Rf as Sceletium was noted along with other constituents. Mesembryanthemum cordifolium (syn. Aptenia cordifolia) This has been studied and seems to contain 4,5-dihydro-4’-O-methylsceletenone and 4’-O-methylsceletenone along with flavanoids, tannins, phenols, saponins, and cinnamic acid derivatives (and esters) The characteristic Sceletium constituent was absent in these but other different compounds were present. Will follow up with more conventional silica TLC which will hopefully have more easily reproducible results Edited April 1, 2023 by Alchemica 3 2 Share this post Link to post Share on other sites
Alchemica Posted March 30, 2023 (edited) Citizen Plant Science - Exploring the Aizoaceae Despite a long history of traditional use as medicines, the Aizoaceae remain an under-studied family of plants [1]. They often contain phytocompounds such as alkaloids, flavonoids, steroids, and their related intermediary compounds, one of the commonly found alkaloids is the mesembrine-type alkaloids [2,3]. These plants are well known in traditional medicine systems for antidepressant, anxiolytic, antimicrobial, anti-inflammatory, and antioxidant effects. Sceletium tortuosum (Figure 1.) contains mesembrine and derivatives (Figure 2.), an alkaloid not typically found in the emarcidum-type Sceletium spp. Other plants such as Delosperma pruinosum, and Delosperma pottsii [2] appeared to have some level of mesembrine but while Mesembryanthemum cordifolium syn. Aptenia cordifolia was initially thought to contain some levels of mesembrine, it seems instead to contain derivatives such as mesembrane sceletenone derivatives [4] which have in initial studies shown antidepressant activity superior to imipramine [5]. Figure 1. S. tortuosum Figure 2. Mesembrine and derivatives. Other Sceletium spp. have been traditionally used, S. emarcidum "was valued as highly as S. tortuosum in Southern Africa by different tribes and makes a very good Kougoed product". While apparently devoid of mesembrine, it seems to contain 4'-O-demethylated mesembrine-type alkaloids [6]. Mesembrine has been found in Amaryllidaceae such as Narcissus pallidulus and Narcissus triandrus. Other Mesembryanthemums such as M. crystallinum and M. nodiflorum have been suggested to contain some levels of mesembrine. More recently, Lampranthus spp. (Figure 3.) have been specifically marketed as Sceletium substitutes 'Chinese Kanna', or used as adulterants. These seem to contain mesembrenol and low levels of mesembrenone. Figure 3. Lampranthus and mesembrenol One early explorer called S. tortuosum ‘the greatest Clearer of the Spirits, and the noblest Restorative in the World’. Traditional use of Sceletium in South Africa often involves the collection and fermentation of the plant material, which has been studied and lead to greater levels of mesembrine and changes to levels of mesembrenone [7] and reduce oxalate levels. Mesembrine is a potent serotonin reuptake inhibitor (Ki = 1.4 nM) and to a lesser extent PDE4 inhibitor. Other notable mesembrine-type alkaloids include mesembrenone (also a potent serotonin reuptake inhibitor [Ki = 27 nM] and a more potent PDE4 inhibitor than mesembrine itself), and mesembranol. Another assay found that Sceletium tortuosum comprising of > 70% (w/w) stabilized mesembrine was a monoamine releasing agent [8]. The pharmacological properties of mesembrine and it's derivatives are quite diverse, showing anti-inflammatory, cytoprotective, VMAT-2 upregulation, mild inhibition of AChE, mild MAO inhibition, limited reuptake of NE and DA at high concentrations. Mesembrenol and mesembrenone seem to also share the SERT inhibitory and PDE4 inhibitory activitty. Clinically, the Sceletium spp. have shown antidepressant, anxiolytic and cognitive enhancement effects, case studies demonstrating increased attention, focus, and motivation while a patient with “a baseline mood of depression alternating with anxiety” felt “more focused, more engaged, and not so socially distant” after ten days [9]. Species such as Delosperma bosseranum (Figure 4) have been suggested by members of the ethnobotanical community to offer similar effects to Sceletium spp. but remain uncharacterised phytochemically and pharmacologically. Others such as Trichodiadema spp. (Figure 4), traditionally used as beer making roots known as 'Karee Moer' (T. barbartum syn. stellatum) as suggested to contain mesembrine but a detailed analysis is lacking (limited to positive alkaloid tests). Figure 4. The prototypical mesembrine containing plant, Sceletium and D. bosseranum and T. barbatum The purpose of this citizen science was to explore a diverse range of Aizoaceae for mesembrine-type alkaloids, including uncharacterised ones such as Delosperma bosseranum and Trichodiadema barbatum and screen diverse Mesembryanthemums for possible mesembrine-type alkaloids. [1] https://doi.org/10.1016/j.jep.2022.115988 [2] https://doi.org/10.1016/j.phytochem.2019.112061 [3] https://doi.org/10.1076%2Fphbi.36.3.173.6350 [4] https://def-sa.com/wp-content/uploads/2020/08/Gaffney_Candice_D_2006_0.pdf [5] https://doi.org/10.1080/14786419.2020.1788019 [6] https://doi.org/10.3389%2Ffnut.2022.819753 [7] https://doi.org/10.1016/j.sajb.2018.10.011 [8] https://patents.google.com/patent/US20160038551A1/en [9] https://www.researchgate.net/publication/328942189_Kabbo's_Kwain_The_Past_Present_and_Possible_Future_of_Kanna A range of Aizoaceae (Silica, I2 visualisation) gave diverse phytochemical profiles Trichodiadema stellatum (root) TS Trichodiadema bulbosum (leaf/stem) TB Sceletium tortuosum (root) ST Sceletium emarcidum (leaf/stem) SE Aptenia cordifolia AC ? Unknown Mesemb. Drosanthemum floribundum Dros. Mesembryanthemum (Red) [?Lampranthus] MR Mesembryanthemum (Gold) MG Mesembryanthemum (Yellow, squarish stems) MY Mesembryanthemum (White) MW D. bosseranum DB D. floribundum DF 1. acetone elution 2. acetone:white spirits:1:1 elution To TLC soon Tetragonia tetragonioides Mesembryanthemum crystallinum Lampranthus spectabilis (Red) D. echinatum Edited April 3, 2023 by Alchemica 2 2 Share this post Link to post Share on other sites
Darklight Posted March 30, 2023 Morning lovely human. @Alchemica I so admire your work, you're a champion mate and that's an understatement <3 Beautiful work, thank you for posting it here Would it be possible to get reference compounds for molecules of interest? I know they're hell exxy, but is it possible? Or can you get reliability via the rf without them? 2 Share this post Link to post Share on other sites
El Presidente Hillbillios Posted March 31, 2023 i really think there is a lot to learn with these plants. thanks for putting in the effort for us the Trichodiadema that resides at my place is really starting to crank, i look forward to distributing it around to the local plant people 2 Share this post Link to post Share on other sites
Alchemica Posted April 11, 2023 After some peer scrutiny on the initial attempt and revisions, here is a rough draft. mesemb paper V2.pdf 3 2 Share this post Link to post Share on other sites
Darklight Posted April 12, 2023 On 11/04/2023 at 12:23 PM, Alchemica said: After some peer scrutiny on the initial attempt and revisions, here is a rough draft. mesemb paper V2.pdf How awesome this is makes me really happy Lovely work my friend. So much happiness for its beauty If youse haven't already read it, can definitely recommend Thank you for sharing Share this post Link to post Share on other sites
Darklight Posted April 12, 2023 On 01/04/2023 at 10:07 AM, El Presidente Hillbillios said: i really think there is a lot to learn with these plants. thanks for putting in the effort for us the Trichodiadema that resides at my place is really starting to crank, i look forward to distributing it around to the local plant people Yeah mine is taking off too, given how soggy and mouldy the local climate can be here I thought it would be doomed Na, it's loving life, third set of new leaves I think Hillbilly, be good to do side by side TLCs to compare climate, season and propagation mix differences over time to complement Alchemica's work, what do you reckon? Share this post Link to post Share on other sites
El Presidente Hillbillios Posted April 12, 2023 yeah i think thats an awesome idea! Share this post Link to post Share on other sites
Freakosystem Posted April 15, 2023 Nice to see a mention of one of our native species; Tetragonum tetragonioides. I was very surprised to see it growing in almost every vegie garden I spotted in France a few years ago. It seems a lot more popular there than here, although always exclusively for edible qualities. I've never heard of any antidepressant effects from it previously. 1 Share this post Link to post Share on other sites
fyzygy Posted April 15, 2023 16 hours ago, Freakosystem said: It seems a lot more popular there than here Like a lot of these plants, T. tetragonioides makes an excellent ground cover. I don't know why it's such an uphill battle persuading people to cook it and eat it. I think people are (literally) sold on supermarket varieties of fresh food. As for "weeds," even if they are superfoods ... forget it. Definitely needs cooking but, and I wonder if that has an effect on medicinal value? 1 Share this post Link to post Share on other sites
Alchemica Posted April 16, 2023 (edited) Thanks for the input, feedback and for reading Just random bits of non-Aizoaceae TLC info whilst experimenting Tabernaemontana undulata (root bark) and T. orientalis (stem) Samples were macerated in basified isopropanol (aq. ammonia), decanted and concentrated. TLC (silica, glass backed, 0.2mm, I2 visualisation) with acetone elution gave significant trailing and poor separation of the constituents for both. Elution with acetone:white spirits 1:1 gave some separations of compounds from T. undulata (Rf = 0.13, 0.35, 0.56, 0.89) but poor separation of constituents in the T. orientalis.(one potential constituent Rf = 0.75). White spirits elution alone gave a band of constituents ~ Rf = 0.24 for both Heimia Contains biphenylquinolizidine lactone alkaloids [1]. Plant material (leaf/stem) was soaked in basified isopropanol (aq. ammonia) and concentrated. Initial elution with acetone gave poor results but TLC (silica, 0.2mm, acetone:white spirits 1:1) gave distinct separation of compounds Rf = 0.43, 0.48, 0.55, 0.63, 0.95 Two (Rf = 0.43 and 0.95) were green-yellow/ yellow and fluorescent, the others plain visible pigments (green, blue-grey) [1] https://www.shaman-australis.com/~auxin/heimia.html Lobelias TLC with these posed issues in finding an adequate solvent system. L. cardinalis: Lobinaline as the major alkaloidal constituent (below) L. tupa: lobelanidine, lobeline and pentylsedinine etc (below) Dried aerial plant material was allowed to sit overnight in basified isopropanol (aq. ammonia), decanted and concentrated. TLC (silica, glass backed 0.2mm, acetone elution, I2 visualisation) gave a distinct compound Rf = 0.65 for the L. cardinalis but did not give any notable visible constituents for the L. tupa. TLC (silica, glass backed 0.2mm, acetone:white spirits 1:1, I2 visualisation) gave a distinct compound Rf = 0.40 for L. cardinalis and hints of a potential compound Rf = 0.31 for the L. tupa. Ashwagandha leaf withanolides Leaf withanolide constituents mainly include withaferin A and withanone Crushed dried leaf was macerated in isopropanol and the sample concentrated. TLC (silica, glass backed, 0.2 mm, I2 visualisation) gave poor results eluted with acetone but with mixed solvent (acetone:white spirits 1:1) gave at least four constituents Rf = 0.23, 0.33, 0.48 and 0.55 Exploring a relatively uncharacterised Leguminosae - Senegalia rugata Senegalia rugata syn. A. concinna - Sompoi or Shikakai - seen in Fig. 1 and 2. belongs to the Leguminosae family and is a medicinal plant widely grown in Southern Asia including India, Myanmar and Thailand. It is commonly known as shikakai and soap pod as it is used for cleansing of hair as a natural shampoo. The phytochemistry has been explored only to a limited extent [1,2]. There is also suggestion of internal use and the calyctomine (Fig. 3) has been used in the supplement industry [3]. Initial phytochemical study Figure 3. calyctomine Alkaloids, flavonoids, saponins, and phenolics were found in all extracts while phytosterols was found only in the ethanol extract. It yields lupeol, spinasterol, acacic acid, lactone, and the natural sugars glucose, arabinose and rhamnose. It also contains hexacosanol, spinasterone, oxalic acid, tartaric acid, citric acid, succinic acid, ascorbic acid, and the alkaloids calyctomine, a tetrahydroisoquinoline, and nicotine. Various other compounds have been isolated and characterised from the pods of this plant such as epigallocatechin, acacic acid lactone, machaerinic acid and its lactone, sapogenin-B, acacidiol, acacigenin-B, and kinmoonosides Experimental: Plant material was macerated in basified isopropanol (NaOH) and concentrated. Initial TLC (Silica, glass backed, 0.2mm, acetone elution, I2 visualisation) gave two main constituents, Rf = 0.14 and 0.59 but there was significant trailing of the compounds. Elution with mixed solvent (acetone:white spirits 1:1) gave a major constituent Rf = 0.65. [1] Screening of Secondary Metabolites and Antibacterial Activity of Acacia concinna http://dx.doi.org/10.3923/jm.2010.974.979 [2] Isolation and Evaluation of Anti-inflammatory activity of Epigallocatechin from Senegalia rugata along with PUFAs https://rjptonline.org/AbstractView.aspx?PID=2021-14-11-23 [3] Calyctomine HCL? https://anabolicminds.com/community/threads/calyctomine-hcl.282282/ Edited April 16, 2023 by Alchemica 1 1 Share this post Link to post Share on other sites
Darklight Posted April 19, 2023 I can't keep up with you mate, I'll have to digest this later. Still going through your lovely Azo paper and looking at replicating it alongside some new variables here nom...nom nom :D Such lovely work, thank you Definitely needs cooking but, and I wonder if that has an effect on medicinal value? Yeah I've been wondering the same. If there are easily made anni-d extracts that bypass the oxalates that'd be cool tho. I'd still like to know the difference between cooked and raw for potential supplementation to the same effect tho- I'd rather cook a feed occasionally than harvest a bunch to extract Share this post Link to post Share on other sites
Alchemica Posted June 8, 2023 (edited) I've been busy expanding on this research, hopefully it's of interest/inspiration to others: So far the research extends to: Citizen TLC Phytochemical Screening of Diverse Mesembryanthemums https://pdfhost.io/v/gugGrtnCj_ Phytochemistry of Aptenia https://pdfhost.io/v/dq6SpR9nM_aptenia Citizen TLC analysis of marketed commercial Sceletium tortuosum products: https://pdfhost.io/v/SVvI3PQwb_TLC_commercial_sceletium_produc Using TLC to guide discovery of hypothetical new Kanna substitutes https://pdfhost.io/v/A4u.08qVQ_kanna_substitutes_TLC Utilising yeasts to encourage the bioconversion of mesembrine-type alkaloids yeasts to encourage bioconversi | PDF Host Initial TLC study of an uncharacterised Sceletium ('Little Karoo') https://pdfhost.io/v/.ASjnTMdo_Initial_TLC_study_of_an_unchara Sceletium Chemotypes - Characterisation of a 'Superior Quality' Sceletium product sceletium chemotypes | PDF Host Reagent analysis of phytochemicals - application to Sceletium spp and Mesembryanthemums. Reagent analysis of phytochemicals - application to Sceletium spp and Mesembryanthemums. | PDF Host Edited October 26, 2023 by Alchemica 2 1 Share this post Link to post Share on other sites
