Journal of
Medicinal Plants Research

  • Abbreviation: J. Med. Plants Res.
  • Language: English
  • ISSN: 1996-0875
  • DOI: 10.5897/JMPR
  • Start Year: 2007
  • Published Articles: 3684

Full Length Research Paper

Acute toxicity and histopathological assessment of methanol extract of Cleome viscosa (Linn.) whole plant.

Taiwo O. Elufioye*
  • Taiwo O. Elufioye*
  • Department of Pharmacognosy, Faculty of Pharmacy, University of Ibadan, Nigeria.
  • Google Scholar
Joel O. Onoja
  • Joel O. Onoja
  • Department of Pharmacognosy, Faculty of Pharmacy, University of Ibadan, Nigeria.
  • Google Scholar


  •  Received: 03 February 2015
  •  Accepted: 16 March 2015
  •  Published: 17 March 2015

 ABSTRACT

Cleome viscosa Linn (Cleomaceae) is a medicinal plant used widely in Nigeria for the management of various ailments.  This research appraised the toxic potential of the plant with a view to validating or contesting its safety. Acute oral toxicity of the methanolic whole plant extract of Cleome viscosa was evaluated in mice using modified Lorke’s method. Signs accompanying toxicity and possible death of animals were investigated for a period of two weeks to determine the median lethal dose (LD50) of the extract. After two weeks observation period, all the animals in the respective dose groups 10, 100, 1000, 1600, 2900 and 5000 mg/kg were euthanized by cervical dislocation. The weight gained, absolute organ weight, and mean organ-body weight ratios (OBR) were determined and compared with values from those of the control group. The oral median lethal dose of the extract was found to be greater than 5000 mg/kg. There was a significant difference in weight gained on day 7 (P=0.052) among dose groups up to 1000 mg/Kg body weight. There was however, no significant difference in the relative organ weights between treated and control animals except for the Liver (p=0.048).  Histopathological analysis showed mild congestion of the pulmonary vessels at dose 1600 mg/kg and above, mild diffuse vacuolar degeneration of hepatocytes across all tested dose as well as mild renal cortical congestion especially at high dose. The oral median lethal dose results indicate that the methanol extract of Cleome viscosa whole plant is non- toxic by oral administration at the tested doses.

 

Key words: Cleome viscosa, methanol extract, acute toxicity, histopathology


 INTRODUCTION

For centuries and in most of the cultures throughout the world, herbal prescriptions and natural remedies are commonly employed for relief or treatment of diseases (Maqsood et al., 2010). Also in modern world, herbal medicines are becoming popular as people resort to natural therapies. Novel clinically active drugs are been isolated from higher plants. Regrettably, there are limited scientific evidence as to the efficacy and safety to back up the continued therapeutic application of these medications. The justification for their use has rested largely on long term clinical knowledge (Zhu, 2002).  Now, with the upsurge in the use of herbal medicines, a comprehensive scientific exploration of these plants will go a  long  way  in substantiating  their  folkloric usage as well as their prophylactic properties (Sofowora, 1993). One foremost and prevailing benchmark in the selection of herbal medicines for use in health services is safety. Plants extracts should not only be efficacious but safe for consumption.

 

Cleome viscosa Linn. (Cleomaceae) is a weed distributed throughout the tropical regions of the world and plains of India. The plant is an annual, sticky herb with a strong penetrating odour, yellow flower and long slender pods containing seeds.  In Ayurvedic system of medicine, the plant is used for the treatment of fever, inflammations, liver diseases, bronchitis and diarrhea (Chatterjee et al., 1991). The rural people use the fresh juice of the crushed seed for the treatment of infantile convulsions and mental disorder.  The juice of the plant diluted with water is given internally in small quantities in fever and the leaves are useful in healing wounds and ulcer (Nadkarni, 1982; Kirtikar et al., 1984).

 

The smoke from its leaves is used by the locals to repel mosquitoes at night. Its extract exhibited larvicidal activity against the second and forth instar larvae of Anopheles stephensi, a vector of malaria in India (Saxena et al., 2000). C. viscosa is highly effective in a wide spectrum of diseases and reported to possess antidiarrhoeal (Devi et al., 2002), analgesic (Parimaladevi et al., 2003), antipyretic activity (Devi et al., 2003), psycho-pharmacological, anti-microbial properties including in vitro Helicobacter pylori and wound healing activity (Parimala et al., 2004a; Mahady et al., 2006; Panduraju et al., 2011), also against Escherichia coli , Proteus vulgaris and Pseudomonas aeruginosa (Sudhakar et al., 2006). In view of the reported effects of C. viscosa, the toxic potential of this plant was studied to generate information on its toxicity profile.


 MATERIALS AND METHODS

Plant materials

 

The plant, C. viscosa Linn. was collected from Jeje area of Ibadan, Oyo State and  authenticated at the Forestry Research Institute of Nigeria where voucher specimen was deposited under the reference number FHI 109669. The whole plant was dried at room temperature and powdered. About 2 kg of the powdered sample was soaked with 100% methanol for 48 h. The extract was concentrated using rotary evaporator and percentage yield was 5.12%. The dry extract was stored in a refrigerator at 4°C for further use.

 

 

Animals

 

The animals (ICR mice), both male and female, 6 to 7 weeks old (15 to 27g) used for these experiments were obtained from the Animal House, Department of Zoology, University of Ibadan. The mice were housed under standard conditions, fed with standard animal feed and given water ad libitum throughout the study period. They were allowed to acclimatize for seven days before the test was commenced. All experimental protocols were in compliance with University of Ibadan Ethics  Committee  Guidelines  as  well  as internationally accepted principles for laboratory animal use, and care as found in the US guidelines (NIH publication Number 85-23, revised in 1985).

 

 

Phytochemical screening

 

Preliminary phytochemical screening was carried out according to Harborne, 1998.

 

 

Acute toxicity study

 

Acute toxicity study was carried out according to modified Lorke’s method (Lorke, 1983). The study was conducted in two phases using a total of sixteen animals.  The mice were fasted overnight prior administration of plant extract. In the first phase, twelve animals were divided into 4 groups of 3 mice each. Groups 1, 2 and 3 animals were given single dose of 10, 100 and 1000 mg/kg of the extract orally, respectively, to establish the possible range of doses producing any toxic effect. Group 4, the control group received a mixture of distilled water and dimethyl sulfoxide (DMSO). In the second phase, the first three animals received 1600, 2900 and 5000 mg/kg separately, while the forth (the control) received a mixture of distilled water and DMSO. All animals were observed frequently on the day of treatment and surviving animals were monitored daily for 2 weeks for signs of acute toxicity. Recovery and weight gain were seen as indications of having survived the acute toxicity. The weights of these organs were also taken and the mean organ-body weight ratios calculated and compared with those of the control group.  Body weights of the mice were recorded on study days 0 (initiation), 7 and 14 (termination). At the end of 14 days, all surviving mice were euthanized by cervical dislocation. Five organs, heart, lungs, liver, kidney and spleen were isolated and subjected to complete gross necropsy and histopathological study. Histopathological assessment and photomicrography of prepared slides were done using an Olympus light Microscope with attached Kodak digital camera. % Relative organ weight= Absolute organ weight (g)/Body weight of mice on sacrifice day x100. Figure 1 to 5.

 

 

 

 

 

 

Statistical analysis

 

The statistical analyses were carried out using Statistical Package for Social Sciences (SPSS-17 computer package) and ANOVA (one-way) followed by Duncan’s Multiple Comparism Test. All data were expressed as mean ± SD of triplicate parallel measurements. Differences between means at 5% level (p ≤ 0.05) were considered significant.


 RESULTS AND DISCUSSION

Despite the widespread use of medicinal plants, few scientific studies have been undertaken to ascertain the safety and efficacy of traditional remedies. To determine the safety of drugs and plant products for human use, toxicological evaluation is carried out in various experimental animals to predict toxicity and to provide guidelines for selecting a ‘safe’ dose in humans. The highest overall concordance of toxicity in animals with humans is with hematological, gastrointestinal, and cardiovascular adverse effects (Olson et al., 2000), while certain adverse effects in humans, especially hypersensitivity and idiosyncratic reactions, are poorly correlated with toxicity observed in animals. Furthermore, it is quite difficult to ascertain certain adverse effects in animals such as headache, abdominal pain, dizziness and visual disturbances. In addition, interspecies differences in the pharmacokinetic parameters make it difficult to translate some adverse effects from animals to humans (Olson et al., 2000).  The antipyretic, analgesic, and anti- inflammatory (Parimala et al., 2003a, b) as well as antimicrobial (Sudhakar et al., 2006), psychopharmacological effects (Parimala et al., 2004b) and immunomodulatory effects (Tiwari et al., 2004) of C. viscosa has been reported.

 

The    biological/pharmacological   activity,  as  well  as toxicity potential of a plant is directly related to the type, nature and quantity of secondary metabolites present in it. Thus, screening for the presence of possible phytoche-micals in a plant is imperative. The results of preliminary phytochemical screening are given in Table 1. It shows the presence of flavonoids, phenolic compounds, alkaloids, phytosterol, fatty acid and saponins.  Anthraquinone, tannin and coumarins were absent in the methanolic extract of C. viscosa L.

 

Flavonoids and other phenolics are ubiquitous in nature and can occur either in the free state or as glycosides. They constitute one of the most characteristic classes of compounds in higher plants and many are easily recognized as flower pigments in most flowering plants. However, their occurrence is not restricted to flowers but include all parts of the plant. They are widespread and have relatively low toxicity compared to other active plant compounds.  Flavonoids have potential to be biological "response modifiers", such as anti-allergic, anti-inflammatory, anti-microbial and anti-cancer.

 

Phytosterols also have been implicated in lowering cholesterol (Pollak, 1953; Tilvis and Miettinen, 1986) and inhibiting lungs, breast, ovarian and stomach cancer (Woyengo et al., 2009). They also have long history of safety (Jones 2007). Medicinal use of alkaloid-containing plants has a long history (Hesse, 2002). The percentage of alkaloids in plants is usually small, and is not homogeneous over the plant tissues. Depending on the plants, the maximum concentration could be observed in the leaves fruits, seeds, root or bark (Grinkevich, 1983). Furthermore, different tissues of the same plants may contain different alkaloids (Orekhov, 1955). Consuming some secondary metabolites can have severe con-sequences. Alkaloids can block ion channels (Hamill and McBride, 1996), inhibit enzymes (Pastuszak et al., 1990), or interfere with neurotransmission producing hallucina-tions (Gaudreau and Gagnon, 2005), convulsion, vomiting and even death (Audi, 2005), diterpene gossypol blocks phosphorylation and is very toxic, spinasterol   from   spinach   interferes   animal  hormone actions, gallotannins also binds to protein and block digestion (Hartmann, 2007). Plants containing cyanogenic glycosides can liberate cyanide which blocks cytochrome C-oxidase thus, becoming potentially poisonous (Venturi, 2011). Some phenolics can be carcinogenic while tannic acid has been shown to cause damage to intestinal walls (Glenn, 2005). Saponins are known to have deleterious haemolyzing effect on circulating erythrocytes (Sofowora, 1993).

 

 

 

 

 

 

 

The acute lethal study of C. viscosa on mice (Table 5) showed that no animal died within 24 h after oral administration of the extract, and the LD50 was greater than 5000 mg/kg. The major signs of toxicity noticed within 24 h include ataxia, lethargy and asthenia.  These signs were not seen in 10 mg/kg dose group but progressed and became increasingly pronounced as the dose increased towards 5000 mg/kg b.w.  The LD50, being greater than 5000 mg/kg b.w., is thought to be safe as suggested by Lorke (Tijani et al., 1986; Deora et. al., 2010). Again, the absence of death among mice in all the dose groups throughout the two weeks of the experiment seems to support this claim. The LD50 value of more than 5,000 mg/kg, showed that the extract is practically safe.  

 

Also in the toxicity studies, mice in all experimental group gained weight over the course of this study (Table 2 to 5). There was a significant difference in body weight gained on day 7 (p>0.052) among dose groups up to 1000 mg/Kg body weight. Mice in all experimental group gained weight over the course of this study especially those mice that took higher doses (Table 3). There was however no statistically significant differences (p>0.05) noted in absolute organ weights between the C. viscosa extract treated and control groups.  Also, there was no statistically significant difference in relative organ weights between treated and control animals except  for  the  liver except for the Liver (p=0.048). Liver weight relative to body weights increased in a dose dependent manner in all group with the test extract (Table 4) with the highest liver weights at dose 2900 mg/kg body weight. However, the magnitudes of the alterations were small and were not considered treatment-related. Mild diffuse vacuolar degeneration of hepatocytes and moderate portal congestion of the liver appears to be the major gross pathology accompanying treatment of mice with methanolic with methanolic extract of C. viscosa (Table 6). Again, liver congestion could be attributed in part to its role in biotransformation of xenobiotics or to a slight clog of liver which is a function of lipid metabolization at that dose apart from vascular changes which could be attributed to the treatment.

 

The findings of this study indicate that the methanolic whole plant extract of C. viscosa may be considered safe for consumption since no animal died within 24 h after oral administration of the extract and the LD50 was greater than 5000 mg/kg.


 CONCLUSION

The methanol extract of Cleome viscosa whole plant appears non- toxic by oral administration at the tested doses as indicated by the high oral median lethal dose.


 CONFLICT OF INTEREST

The authors declare that they have no conflicts of interest.



 REFERENCES

Audi J, Belson M, Patel M, Schier J, Osterloh J (2005). Ricin poisoning: a comprehensive review. JAMA 294(18):2342-2351.
Crossref
 
Chatterjee A, Prakashi SC (1991). The Treatise on Indian Medicinal Plants, 2nd Eds., Vol. I. New Delhi, Council for Scientific and Industrial Research. P 155.
 
Deora PS, Mishra CK, Mavani P, Asha R, Shrivaatava B, Rajesh KN (2010). Effective alternative methods of LD50 help save numbers of experimental animals. J. Chem. Pharm. Res. 2(6):450-453.
 
Devi BP, Boominathan R, Mandal SC (2002). Evaluation of anti-diarrheal activity of Cleome viscosa L. extract in rats. Phytomedicine 9(8):739-742.
Crossref
 
Devi BP, Boominathan R, Mandal SC (2003). Evaluation of antipyretic potential of Cleome viscosa Linn. (Capparidaceae) extract in rats. J. Ethnopharmacol. 87(1):11-13.
Crossref
 
Gaudreau JD, Gagnon P (2005). Psychotogenic drugs and delirium pathogenesis: the central role of the thalamus. Med. Hypotheses 64(3):471-475.
Crossref
 
Glenn I (2005). The role of plant secondary metabolites in mammalian herbivory: ecological perspective. Proc. Nutr. Soc. 64:123-131.
Crossref
 
Grinkevich NI, Safronich LN (1983). The chemical analysis of medicinal plants. Proceedings Allowance Pharmaceutical Universities. pp. 122-123.
 
Hamill OP, McBride DW (1996). The pharmacology of mechanogated membrane ion channels. Pharmacol. Rev. 48(2):231-252.
Pubmed
 
Harborne JB (1998). A Guide to modern techniques of plant Analysis. USA: Kluwer Academic Publisher.
 
Hartmann T (2007). From waste products to ecochemicals: fifty years research of plant secondary metabolism. Phtyochemistry 68:22-24.
Crossref
 
Hesse M (2002). Alkaloids: Nature's Curse or Blessing? Wiley VCH. P 303.
 
Jones PJ (2007). Ingestion of phytosterols is not potentially hazardous. J. Nutr. 137(11):2485.
Pubmed
 
Kirtikar KR, Basu BD (1984). Indian Medicinal plants, Lalit Mohan Basu Publication, Dehra Dun, 2nd Edn, vol. 1. pp. 181-185.
 
Lorke D (1983). A new approach to practical acute toxicity testing. Arch. Toxicol. 53:275-287.
Crossref
 
Mahady GB, Bhamarapravati S, Adeniyi BA, Penland SL, Doyle B, Locklear T, Solver C (2006). Traditional Thai Medicines inhibit Helicobacter pylori in-vitro and in-vivo: support for ethnomedical use. Ethnobot. Res. Appl. 4:159-165.
 
Maqsood S, Singh P, Samoon MH, Balange AK (2010). Effect of dietary chitosan on non-specific immune response and growth of Cyprinus carpio challenged with Aeromonas hydrophila. Inter Aqua Res. 2:77-85.
 
Nadkarni AK (1982). Indian Materia Medica. Popular Prakashan Bombay. 1:351-352, 498.
 
Olson H, Betton G, Robinson D, Thomas K, Monro A, Kolaja G, Lilly P, Sanders Sipes G, Bracken W, Dorato M, Deun KV, Smith P, Berger B, Heller A (2000). Concordance of toxicity of pharmaceuticals in humans and in animals. Regul. Toxicol. Pharmacol. 32:56-67.
Crossref
 
Orekhov AP (1955). Chemistry of alkaloids (Acad. 2 ed.). M.: USSR. P 12
 
Panduraju T, Parvathi B, Rammohan M, Srinivas RC (2011). Wound healing properties of Cleome viscosa Linn. Hygeia J. Drugs Med. 3:41-45.
 
Parimala B, Boominathan R, Mandal SC (2003a). Evaluation of anti-inflammatory activity of Cleome viscosa. Indian J. Nat. Prod. 19:8-12.
 
Parimala DB, Boominathan R, Mandal SC (2003b). Studies on anal-gesic activity of Cleome viscosa in mice. Fitoterapia 74(3):262-266.
Crossref
 
Parimala DB, Boominathan R, Mandal SC (2004a). Studies on psychopharmacological effects of Cleome viscosa Linn. extract in rats and mice. Phytother. Res. 18:169-172.
Crossref
 
Parimala DB, Boominathan R, Mandal SC (2004b). Studies on psychopharmacological effects of Cleome viscosa Linn. extract in rats and mice. Phytother. Res. 18:169-172.
Crossref
 
Parimaladevi B, Boominathan R, Mandal SC (2003). Studies on anal-gesic activity of Cleome viscosa in mice. Fitoterapia 74(3):262-266.
Crossref
 
Pastuszak I, Molyneux RJ, James LF, Elbein AD (1990). Lentiginosine, a dihydroxyindolizidine alkaloid that inhibits amyloglucosidase. Biochemistry 29(7):1886-1891.
Crossref
 
Pollak OJ (1953). Reduction of blood cholesterol in man. Circulation 7(5):702-6.
Crossref
 
Saxena BR, Koli MC, Saxena RC (2000). Preliminary ethnomedical and phytochemical study of Cleome viscosa. Ethnobotany 12:47-50.
 
Sofowora AE (1993). Medicinal plants and traditional medicine in Africa. Spectrum Books Limited, Ibadan, Nigeria. pp. 150-156.
 
Sudhakar M, Rao CV, Rao PM, Raju DB (2006). Evaluation of antimicrobial activity of Cleome viscosa and Gmelina asiatica. Fitoterapia 77(1):47-49.
Crossref
 
Tijani A, Pomtantz A, Khawar R, Fusi M (1986). Community acquired infections in children on maintenance cyclosporine therapy. Int. J. Pediatr. Nephrol. 7:141-144.
 
Tilvis RS, Miettinen TA (1986). Serum plant sterols and their relation to cholesterol absorption. Am. J. Clin. Nutr. 43(1):92-7.
Pubmed
 
Tiwari U, Rastogi B, Thakur S, Jain S, Jain NK (2004). Studies on the immunomodulatory effects of Cleome viscosa. Indian J. Pharm. Sci. 66:171-176.
 
United Nations (2005). Globally Harmonized System of Classification and Labelling of Chemicals (GHS), 1st ed.; United Nations: Geneva, Switzerland.
 
Venturi S (2011). Evolutionary significance of iodine. Curr. Chem. Biol. 5(3):155-162.
Crossref
 
Woyengo TA, Ramprasath VR, Jones PJH (2009). Anticancer effects of phytosterols. Eur. J. Clin. Nutr. 63(7):813-20.
Crossref
 
Zhu M, Lew KT, Leung P (2002). Protective Effects of Plant formula on Ethanol-induced Gastric Lesions in Rats. Phytother. Res. 16:276-280.
Crossref

 




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