Catha Edulis, (vahl) Forsk, (Miraa): Occurrence, Active Constituents And Pharmacological Activity.

Abstract

The Catha edulis material used in this research, unless otherwise stated was obtained as required from River Road, Nairobi between 7.00 a.m. and 9.00. a.m when the material is in its freshest condition. Usually the material had been transported from Meru, approximately 260 Kilometres from Nairobi less than 20 hours before then. Samples of Catha material, weighing 25 grammes were extracted by refluxing with 75 percent ethylalcohol followed by solvent-solvent extraction at PH-5-6 (to remove neutrals and acids) and PH9 (to recover basic substances) using chloroform. The final basic residue was taken up in chloroform and used in subsequent analysis • The best solvent systems for thin 1ayer chromatography (T.l.C) were found to be Ethyl acetate: Nethanol: Ammonia (17:2:1) and Cyclohexane: Chloroform: Diethylamine (5:4:1). Thin layer , Chromatographic plates were coated with silica gel. After development of plotes, cathinone, DNE were viewed under short and long ultraviolet light using uv chromatoview and subsequently sprayed with 0.3% ninhydrin. Cathinone was easily detected under short (UV) light but d-norpseudoephadrine (DNE) was more sensitive to ninhydrin spray reagent than UV light. Samples of Catha edulis plant used for investigating the relative distribution were obtained from Runyenjes (Embu) and Kangeta (Meru) and were preserved in dry ice until analysed two days later. Results obtained with thin layer chromatographic (TLC) and Gas liquid chromatographic (GLC) analysis of samples obtained from two different geographical localities indicated that there is no significant difference in the chemical constituents of Catha material from the two localities DNE was present in all parts of the plant examined except the root. However the concentrations of ONE in the different plant parts varied, with the highest concentration being in the young growing shoots and young leaves and the lowest concentration in the old branches and stem. Cathinone was more restricted than ONE, with high concentrations occurring in the young growing shoots, lesser amounts in the growing branch lets and none in any other plant part. This finding suggests that cathinone is possibly formed first and then converted to the more stable DNE. Individual preference for the freshest and youngest Catha shoots, is well recognized, However in absence of a reliable method for quantitative estimation of cathinone and DNE it is difficult to explain this observation on the basis of relative concentrations of the active constituents in the plant. Results obtained in the present work however show that cathinone and DNE were still present in large amounts in Catha material dried at room Gonditions (20-230c) over a period of 7-10 days. This suggests that t h e po tancv of the Catha material sold in the market (usually wrapped in moist green banana Ieaves) is unlikely to change significantly over a period of even 3-4 days, a suggestion corroborated by some indigenous people who chew Catha material. It therefore appears that the preference for fresh Catha material is based on tenderness palatability and colour appeal rather than on potency. Salts of cathinone and DNE were found to be stable to when dissolved in methanol, dilute ethanol Dr dilute hydrochloric acid. When the salt solutions were kept at ordinary room conditions (20-230c) in colour less glass containers for one month, TLC analysis revealed some degradation. This degradation was even faster with the basic residue from Catha material dissolved in chloroform. Analysis revealed that some cathinone was converted to DNE and one other unindentified substance. No cathinone was detectable in the d-norpseudoephedrine solution which had sparingly changed to another unindentified substance. Cathinone and DNE stored at temperatures below 1oc in solid form, in acidic, basic or neutral solutions or in untreated plant material showed a high degree of stability. Preliminary observations made on whole animals (mice, average weight 25 gms) injected with SO-2oomg/kg cathinone or DNE intraperitonially revealed that these two substances caused central nervous system (CNS) stimulation characterized by hyperactivity, restlessness, hyperresponsiveness, irritability, increased depth and rate of respiration, convulsion (shaking head and body and jumping). This was followed by muscle incordination, respiratory depression (characterised by gasping for air), loss of .righting reflex and for doses equal to or above 200 mg/kg, the animals died within 20-30 minutes. Death could result from circulatory depression and/or respiratory depression. Results obtained from the anaesthetised rat blood pressure showed that cathinone at all concentrations employed caused a sustained fall in blood pressure that lasted even for 3 hours with animals usually dying in the end. DNE on the other hand caused a rise in blood pressure with the first dose, but a second dose always gave ~ fall in blood pressure. This' could be due to cumulation of the drug a point supported by the slow metabolism and excretion of DNE or due to vasodilator effect of DNE as reported in literature. Ephedrine, the prototype of the indirectly acting phenylalkylamines on the same dose range as DNE caused rise in blood pressure and repeated doses gave rise to development of tachyphylaxis. In the anaesthetised rat cathinone and DNE have a different mechanism of action to that of the structural analogue ephedrine. On the isolated rabbit heart cathinone and DNE caused continous depression of the heart at all concentrations employed (10 -2M-lo -7~) with little or no change in chronotropic response. There was no tachyplylaxis evident after repeated dosage. Pretreatment of the rabbit heart with atropine (3xlO-4M) did not antagonise the carciodepressant effect of cathinone and DNE indicating that the~2ffect might not be mediated through release of acetylcholine. Pretreatment of the rabbits with reserpine lmg/rabbit for 3 consecutive days and consequent use of the respective isolated hearts did not alter the response of the isolated heart to cathinone and d-norpseudoephedrine. This is an indication that the drugs have no adranerqi cally mediated mechanism and may be generally depressing the contractile mechanisms in contrasv with ephedrine that potrays chololinergic and adrenergic effects that are governed by dose. Using isolated rabbit aortic strip, a vascular smooth muscle, it was shown that cathinone and d-norpseudoephedrine appreciably antagonised the potassium (K+) induced contractions of the preparation, but ephedrine enhanced the potassium induced contractions. Evidence from literature shows that K+ induced contractions of the rabbit aortic strip is due to transmembrane influx of calcium ions. This partly explains the cardiodepression and hypotension obtained with cathinone and DNE. Results obtained using isolated guinea pig ileum stimulated with barium ions (8a2+) and ONE antagonised the (2xlO-3) re.veale.d that cathinone 8a2+ induced contractions. Cathinone, DNE and ephedrine have a relaxant effect on the electrically stimulated innervated rabbit intestine, the blocked by pretreatment with adrenergic neurone blocker, guanethidine O.lmg/ml bath concentration. Acetylcholine however caused contraction of the smooth muscle in presence of guanethidine, cathinone or ONE indicating that the cholinergic receptors were not affect ed , Preliminary results on whole animals suggested that death of the animals could be due to respiratory depression an effect that could be mediated through the neuromuscular junction, or through paralysis of the nerve or direct depression of the diaphragm muscle. Results obtained with the phrenic-nerve diaphragm preparation showed that cathinone, DNE and ephedrine caused a continous 'depression of the contractile response of the preparation. This blocakade was enhanced in magnitude by lignocaine and d-tubocurarine. The neuromuscular facilitatory action of physostigmine was also blocked by DNE, cathinone and ephedrine. When tested on the directly stimulated diaphragm muscle, cathinone, DNE and ephedrine caused a depression of the contractile response of the muscle. Results obtained with the guinea pig wheal and the frog sciatic nerve preparations, showed that cathinone, DNE and ephedrine had some local anesthetic:activity. Throughout this research work, no adrenergic or cholinergic mediated actions of cathinone or DNE have been indicated. A direct action is implicated. This direct action is thought to be a generalised depression of the contractile tissue, an action usually associated with local anaesthetics and calcium antagonists. In this work cathinone and DNE have been shown to possess some local anaesthetic activity and some calcium antagonistic action. It is therefore possible that the pharmacological actions of these compounds are due to these two properties. Metabolic studies, using urine collected from 4 human volunteers who had ingested 15mg cathinone each, revealed that cathinone is metabolised to DNE and excreted in urine. No cathinone could be detected in the urine collected 15 hours after ingestion while DNE could be detected even after 46 hours. Analysis of urine collected from human volunteers who had taken 15mg DNE showed that DNE is excreated unchanged and no cathinone could be detected at all. Similar metabolic results were obtained in rabbits fed on cathinone and DNE