South African Journal of Botany 2003, 69(2): 144–147 Copyright © NISC Pty Ltd Printed in South Africa — All rights reserved SOUTH AFRICAN JOURNAL OF BOTANY ISSN 0254–6299

The ethnobotany and chemistry of the genus Clivia (Amaryllidaceae)

NR Crouch*1,2, DA Mulholland1, TL Pohl1 and E Ndlovu1,3

1 Natural Products Research Group, Department of Chemistry, University of Natal, Durban 4041, South Africa 2 Ethnobotany Unit, National Botanical Institute, PO Box 52099, Berea Road 4007, South Africa 3 Kwamalulekoes Nursery and Herbalist, PO Box 11409, Mooi River 3300, South Africa * Corresponding author, e-mail: [email protected]

Received 4 June 2002, accepted in revised form 16 October 2002

The herbaceous genus Clivia Lindl. (Amaryllidaceae) thamine, (11S)-11-hydroxyvittatine, lycorine and ster- comprises four species, all of which are employed in bergine were isolated from C. caulescens, and haeman- traditional medicine. Two lesser-known ethnomedicinal thamine, haemanthidine in its free base and quaternary members, C. gardenii Hook. and C. caulescens R.A.Dyer salt form, and the quaternary salt of tazettine from C. are phytochemically characterised, revealing a variety gardenii. The traditional use of the genus Clivia in of isoquinoline alkaloids. Hippeastrine, haeman- southern Africa is overviewed.

Introduction

The genus Clivia (Amaryllidaceae, Haemantheae) is endem- unlikely to have historically been used by the Zulu. In sup- ic to southern Africa and is represented by four species of port of the identification of material as C. gardenii is the perennial evergreen plants found in moist coastal and inland report by Williams et al. (2000) that the common name for forests. These are C. miniata (Lindl.) Regel (2 varieties), C. genus members in the Witwatersrand trade is umayime, in nobilis Lindl., C. caulescens R.A.Dyer and C. gardenii the Zulu vernacular. Additionally, most species (42%) traded Hook., all of which are rhizomatous rather than bulbous at this site are harvested from KwaZulu-Natal (Williams et al. herbs possessing several strap-shaped leaves in a distic- 2000). These authors determined Clivia spp. to be among the hous arrangement. The chemistry of the first two taxa has most commonly available taxa in medicinal plant shops on been investigated previously and a range of isoquinoline the Witwatersrand, with over 70% of traders stocking them. alkaloids have been found. The results of these investiga- Clivia plants play an important role as protective charms: tions have been reviewed by Vildomat et al. (1997). Plants Gerstner (1941a) reported that the Zulu use the roots as a of the Clivia genus are used by traditional healers in South sprinkling charm to ward off evil (intelezi yokuchela) and as Africa in the treatment of a number of ailments of a physio- a love-charm emetic. Species of Clivia are included as logical or spiritual origin, and have been implicated in human chopped components in intelezi mixes traded in the Durban poisonings. Juritz (1915) reported on the death in Willowvale markets (Cunningham 1988). The Zulu name umayime for in the southern Transkei of a sickly infant treated with Clivia Clivia spp. has been variously interpreted as ‘it should stand nobilis. Further investigations by this author revealed that still’ (Williams et al. 1996), or ‘the young women may stay at large quantities of a strong decoction of the plant produced home’ (Gerstner 1941a). In both cases the connotation is of only mildly emetic effects. Veale et al. (1992) considered the plant preventing evil, of halting the course of a disease, ethnomedicines based on C. miniata to be potentially fatal if of stopping or of making to stay (Pujol 1993). An example of taken in large quantities, and noted vomiting, salivation, diar- a protective charm that includes C. miniata as a component rhoea and depression of the CNS as the main poisoning fea- is detailed by Smith and Crouch (1999). tures. All four species are capable of causing dermatitis The large range of isoquinoline alkaloids isolated from (Hutchings et al. 1996). Clivia spp. are generally thought to be responsible for the Reports of trade in C. nobilis plants in the medicinal plant observed bioactivity of the plant extracts. Clivia miniata rhi- markets of KwaZulu-Natal (Cunningham 1988), zome preparations have been recorded as a remedy for Witwatersrand (Williams et al. 2000) and Mpumalanga snakebite (Bryant 1909), specifically for the bite of the puff- (Mander 1997) likely refer rather to C. gardenii in the former adder (Smith 1893), or associated pain (Pujol 1993). Bryant two markets and C. caulescens in the latter. Clivia nobilis (1909) further listed C. miniata as one of the Zulu emetic finds its northernmost distribution immediately to the north of remedies for fevers known as umkhuhlane. The Xhosa use the Kei River (extending south to Alexandria) and so is root decoctions of C. miniata for treating both barrenness South African Journal of Botany 2003, 69: 144–147 145 and urinary complaints (Broster 1982). Clivia caulescens Materials and Methods has also been recorded as ethnomedicinally used by the Swazi, who know the plant as hlakahla-wemahlatsi (Dlamini Plant materials 1981). The material of C. caulescens investigated in the cur- rent phytochemical report was confiscated from an illegal Fresh Clivia caulescens R.A.Dyer rhizomes (2.4kg) were muthi harvester who had collected the plants from near Long collected from Long Tom Pass in Mapumalanga and a Tom Pass in Mpumalanga. voucher lodged for verification purposes (Crouch 758,NH). The leaves of C. miniata are included as a component of Fresh whole plant material of Clivia gardenii Hook. (1.08kg) isihlambezo and inembe mixtures, antenatal tonics adminis- was collected from Ngome, KwaZulu-Natal (Crouch 809, tered with the aim of ensuring healthy foetal growth, or NH). inducing or augmenting labour (Bryant 1909, Gerstner 1941b, Veale et al. 1992, Varga and Veale 1997). The prepa- Extraction and fractionation rations typically take the form of aqueous infusions of the leaves. The uterotonic activity of aqueous extracts of C. Clivia caulescens plant material was macerated and extract- miniata leaves was evaluated using an isolated rat uterus ed with continuous agitation for 72h in ethanol. The gum-like system (Veale et al. 1989). Extracts were shown to possess residue obtained after removal of ethanol was acidified to pH uterine smooth muscle stimulant activity which augmented 4 using concentrated hydrochloric acid. The aqueous mix- the effects of subthreshold and threshold concentrations of ture was extracted with chloroform to produce fraction A oxytocin. Additionally, the Clivia extract elevated the fre- (840mg), The remaining residue was basicified to pH 8 quency of spontaneous uterine contractions. The inclusion using concentrated ammonium hydroxide and extracted with of Clivia preparations in antenatal ethnomedications may chloroform to produce fraction B (3.95g). Thin layer chro- thus be based on such stimulant effects (Veale et al. 1989). matography showed fractions A and B to contain the same Use of such tonics can result in foetal distress due to uterine compounds, so they were combined and separated using hypertonia (Veale et al. 1992), and also an increase in the column chromatography over silica gel (Merck 9385) using a frequency of meconium-stained liquor passage and caesar- dichloromethane/methanol mixture of increasing polarity ian sections (Mabina et al. 1997). Rather surprisingly, the yielding five alkaloids. Clivia gardenii plant material was active uterotonic compound has been found not to be an macerated and extracted in ethanol at room temperature alkaloid but linolenic acid (Sewram et al. 2001). with continuous agitation for 60 hours. After removal of the Compounds isolated previously from C. miniata and C. ethanol using varying ratios of dichloromethane/methanol, nobilis are given in Table 1. The large number of alkaloids, column chromatography (silica gel — Merck 9385) of the including the very unusual alkaloid nobilisine, prompted the gum-like residue yielded three alkaloids. chemical investigation of the remaining two Clivia species.

Table 1: Alkaloids isolated from Clivia

Class of Alkaloid Alkaloid trivial name Clivia miniata* Clivia nobilis* Clivia caulescens Clivia gardenii Lycorine caranine + cliviasindhine + lycorine + + + sternbergine + Homolycorine clivacetine + clivatine + + cliviamartine + cliviasine + cliviasyaline + clividine + clivimine + clivonidine + clivonine + hippeastrine + + miniatine + nobilisine + Haemanthamine haemanthamine + + + haemanthidine +, +** 11-hydroxyvittatine + Tazettine tazettine +** Miscellaneous cliviahaksine + clivialine + clivojuline +

* Vildomat et al. (1997) ** Isolated as an ammonium salt 146 Crouch, Mulholland, Pohl and Ndlovu

Structure determinations 11-hydroxyvittatine, lycorine and sterbergine were isolated, and from C. gardenii, haemanthamine, haemanthidine, in The alkaloids isolated were identified using NMR and MS the free base and quaternary salt form, and the quaternary techniques and the stereochemistry of the 5,10b-ethano- salt of tazettine (Table 1, Figure 1). The free bases of hae- bridge was determined using circular dichroism where nec- manthidine and tazettine were obtained on treatment of the essary. The identification of the alkaloids was confirmed by salts with NaOH. comparison of physical data against literature values. NMR The isolation of the alkaloid lycorine from C. caulescens is spectra were recorded on a Varian Unity Inova 400MHz significant. Lycorine is the most widespread of the

NMR spectrometer in CDCl3 or CD3OD. IR spectra were Amaryllidaceae alkaloids. It is highly toxic (LD50 (dog) 41mg recorded with a Nicolet Impact 400D spectrometer on sodi- kg-1) and frequently the cause of accidental human poison- um chloride plates and calibrated against an air background. ing (especially from consumption of Narcissus species). EIMS were obtained using a Agilent 6890GC with a 5973N Lycorine causes scorbutic symptoms in experimental ani- mass selective-MS system. CD spectra were recorded at mals and acts as a respiratory stimulant. However, this com- the University of the Free State. pound has been shown to have moderate anti-tumour activ- ity, anti-viral properties and to be a weak protozoicide Results and Discussion (Dictionary of Natural Products 2001). Tazettine, here isolat-

ed from C. gardenii, has been shown to have a LD50 (dog) of From C. caulescens, hippeastrine, haemanthamine, (11S)- 71mg kg-1 (Dictionary of Natural Products 2001).

OH OH N Me HO AcO H H H O OH O MeO H H H H O O N N O HO O Lycorinec Sterberginec Hippeastrinec

OCH3 OCH3

OH OH O O

O N O N

OH Haemanthaminec,g Haemanthidineg

OCH3

OH OH c=C. caulescens N CH 3 g=C. gardenii O O OH O N O O

11-Hydroxyvittatinec Tazettineg

Figure 1: Alkaloids from Clivia caulescens and C. gardenii South African Journal of Botany 2003, 69: 144–147 147

Acknowledgements — This research was funded by the NRF and Mabina MH, Pitsoe SB, Moodley J (1997) The effect of traditional the University of Natal Research Fund. We gratefully acknowledge herbal medicines on pregnancy outcome. The King Edward VIII the Wellcome Trust Equipment grant number 052451 for the provi- Hospital experience. South African Medical Journal 87: 1008–1010 sion of the 400MHz NMR spectrometer. We are grateful to Mr Dilip Mander M (1997) Medicinal Plant Marketing and Strategies for Jagjivan and Mr Bret Parel for running NMR and mass spectra and Sustaining the Plant Supply in the Bushbuckridge Area and to Professor Vincent Brandt of the University of the Free State for Mpumalanga Province. DANCED/DWAF, Nelspruit, South Africa use of the CD instrument. The DEAT is acknowledged for funding Pujol J (1993) Natur Africa. The Herbalist Handbook. African Flora MEDBASE, the National Medicinal Plants Database for South Medicinal Plants. Jean Pujol Natural Healers Foundation, Africa. The Mpumalanga Parks Board (Mr M Lötter and Ms S Durban, South Africa Krynauw) and DWAF are thanked for permission to collect samples Sewram V, Raynor, MW, Mulholland, DA and Raidoo, DM (2001) for analysis. Supercritical fluid extraction and analysis of compounds from Clivia miniata for uterotonic activity. Planta Medica 67: 451–455 References Smith AA (1893) A Contribution to South African Materia Medica. Juta, Cape Town, South Africa Broster JA (1982) Amagquirha. Religion, Magic and Medicine in Smith GF, Crouch NR (1999) Rhipsalis baccifera subsp. mauritiana. Transkei. Via Afrika Limited, Goodwood, South Africa Cactaceae. Flowering Plants of Africa 56: 94–98 Bryant AT (1909) Zulu medicine and medicine-men. Annals of the Varga CA, Veale DJH (1997) Isihlambezo: utilization patterns and Natal Museum 2: 1–103 potential health effects of pregnancy-related traditional herbal Cunningham AB (1988) An investigation of the herbal medicinal medicine. Social Science in Medicine 44: 911–924 trade in Natal/Kwazulu. Institute of Natural Resources Report No. Veale DJH, Oliver DW, Arangies NS, Furman KI (1989) Preliminary 29. University of Natal, Pietermaritzburg, South Africa isolated organ studies using an aqueous extract of Clivia miniata Dictionary of Natural Products (2001) 9.2. Chapman Hall, London leaves. Journal of Ethnopharmacology 27: 341–346 Dlamini B (1981) Swaziland Flora: Their Local Names and Uses. Veale DJH, Furman KI, Oliver DW (1992) South African traditional Ministry of Agriculture and Co-operatives, Mbabane, Swaziland herbal medicines used during pregnancy and childbirth. Journal Gerstner J (1941a) A preliminary check list of Zulu names of plants. of Ethnopharmacology 36: 185–191 Bantu Studies 15: 277–301 Vildomat F, Bastida J, Codina C, Nair JJ, Campbell WE (1997) Gerstner J (1941b) A preliminary check list of Zulu names of plants. Alkaloids of the South African Amaryllidaceae. Recent Research Bantu Studies 15: 369–383 and Development in Phytochemistry 1: 131–165 Hutchings A, Scott AH, Lewis G, Cunningham AB (1996) Zulu Williams R, Crouch N, Pettit G (1996) Geophytes of Kwazulu-Natal Medicinal Plants. An Inventory. University of Natal Press, 1. Underworld plants with a special appeal. IBSA Bulletin 44: Pietermaritzburg, South Africa 14–19 Juritz CF (1915) South African plant poisons and their investigation. Williams VL, Balkwill K, Witkowski ETF (2000) Unraveling the com- Report of the 12th Annual Meeting of the South African mercial market for medicinal plants and plant parts on the Association for the Advancement of Science 11: 109–145 Witwatersrand, South Africa. Economic Botany 54: 310–327

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