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Full-Text (PDF) Vol. 9(8), pp. 254-261, 25 February, 2015 DOI: 10.5897/JMPR2014.5497 Article Number: C1D54B651309 ISSN 1996-0875 Journal of Medicinal Plants Research Copyright © 2015 Author(s) retain the copyright of this article http://www.academicjournals.org/JMPR Full Length Research Paper In vitro anti-Salmonella activity of extracts from selected Kenyan medicinal plants Peter Ogoti1*, Esther Magiri1, Gabriel Magoma1, Daniel Kariuki1 and Christine Bii2 1Department of Biochemistry, Jomo Kenyatta University of Agriculture and Technology, P.o Box 62000-00200, Nairobi, Kenya. 2Centre of Microbiology Research (CMR) KEMRI, P.o Box 54840-00200, Kenya. Received 23 June, 2014; Accepted 16 February, 2015 The aim of this study was to determine in vitro anti-Salmonella activity of extracts of five selected Kenyan medicinal plants against Salmonella ser. Typhi and Salmonella ser. Typhimurium. The extracts from Tithonia diversifolia, Warburgia ugandesis, Croton megalocarpus, Carissa edulis and Launae cornuta plants traditionally used in treatment of typhoid fever were screened for anti-Salmonella activity using disc diffusion and microdilution techniques. The results from the present study have shown that out of thirty six extracts investigated, only nine extracts from T. diversifolia and W. ugendensis showed activity against Salmonella ser. Typhi and Salmonella ser. Typhimurium at 1000 mg/ml. The inhibition zone of ethyl acetate, hexane and methanol extracts of T. diversifolia leaves, ethyl acetate and hexane extracts of T. diversifolia flowers, ethyl acetate and hexane extracts of W. ugandensis stem barks, ethyl acetate and hexane extract of W. ugandensis roots ranged from 8 to 18.5 ± 0 mm. These results were comparable with those of ciprofloxacin (19.67 to 26 mm) and chloramphenicol (6.67 to 24.33 mm). The minimum inhibitory concentration (MIC) of the active extracts were in the range of 0.031 to 15.63 mg/ml which compared very well with ciprofloxacin (0.015 to 0.02) and chloramphenicol (0.022 to 0.03 mg/ml). Extracts with anti-Salmonella activity can be used to source antibiotic substances useful in the treatment of typhoid fever. The study provides the scientific basis for the traditional application against typhoid fever. Key words: Anti-Salmonella activity, medicinal plant extracts, minimum inhibitory concentration, disc diffusion technique, microdilution technique, Salmonella strains, typhoid. INTRODUCTION Salmonella serotype Typhimurium (S. ser. Typhimurium), gastroenteritis in millions of people worldwide each year is a Gram-negative bacterial pathogen that infects (Grassl et al., 2008). For instance, the Centre for Disease humans and animals, causing significant morbidity and Control (CDC) estimates that there are nearly 1.4 million mortality worldwide (Fink and Cookson, 2007). It is an food-borne Salmonella infections annually in the USA obligate intracellular bacterial pathogen that causes (Mead et al., 1999). Various strategies have been *Corresponding author. E-mail: [email protected]. Tel: +254700354243, +254728957828. Author(s) agree that this article remain permanently open access under the terms of the Creative Commons Attribution License 4.0 International License Ogoti et al 255 employed in the treatment and management of Kenya Medical Research Institute (CMR-KEMRI) for this study. Salmonella infection. Fluoroquinolones and tetracyclines are most commonly used to treat Salmonella infections. Plant However, Salmonella strains resistant to these antibiotics have been reported in Korea and other countries (Choi et The five plants selected for this study were collected in Nyamira al., 2005; Stevenson et al., 2007). One major concern to County as indicated in Table 1. The plants were authenticated at public health has been the global dissemination of S. Jomo Kenyatta University of Agriculture and Technology, Botany typhimurium Definitive Type 104, which is resistant to Department. cotrimoxazole, nalidixic acid and ampicilin (Perron et al., 2008; Kariuki et al., 2010). The rise in antibiotic-resistant Experimental design strains has led to increased interest in use of plant materials to develop new effective drugs. According to The nine plant parts obtained from 5 selected medicinal plants World Health Organization (WHO) more than 80% of indicated in Table 1 were extracted using four solvent systems world’s population relies on traditional medicine for their namely; hexane, ethyl acetate, methanol and water. The extracts obtained were subjected to standard phytochemical analyses as primary healthcare, majority of who use plant active described by Jigna et al. (2006). In addition, the extracts were principles (Gupta et al., 2005). A wide variety of plants screened for anti-Salmonella activity against four clinical isolates. are used in Africa for treatment of fever, dysentry, Samples in triplicate were subjected to disc diffusion and cholera, diarrhoea and other infections typical of the microdilition in duplicates to confirm anti-Salmonella activity. tropical countries (Ayogu and Amadi, 2009; Ajayi and Akintola, 2010). For instance, traditional practitioners in Preparation of plant materials for extraction Nigeria use herbal preparations to treat microbial infections such as typhoid and paratyphoid infections The plant materials were washed under running tap water and left (Iroha et al., 2010). to drain off. The plant parts were chopped into small pieces. The Plants used in this study have traditionally been dried pieces were ground into powder and their weights recorded. associated with disease curative and preventive practices in many countries for a long time. Garcia and Delgado Extraction techniques (2006) have reported that Tithornia diversifolia has promising medicinal value. Skin products formulated from The nine plant materials were extracted using selected solvents. T. diversifolia extracts have been shown to have Each plant material was extracted sequentially using hexane, ethyl antimicrobial properties (Kareru et al., 2010). acetate and methanol in the order of increasing polarity. Single In Ethiopia Warburgia ugandensis extracts are used to extraction was carried out using water for each of the nine plant materials. treat malaria, tuberculosis, bronchitis, pneumonia, hepatitis, tapeworm, gonorrhea and asthma (Wube et al., 2010; Were et al., 2010; Opiyo et al., 2011). The Preparation of hexane extract decoction from Croton megalocarpus bark is used as a remedy for worms and whooping cough. Grounded roots Approximately 500 g of each plant powder was soaked separately are used for syphilis, anthrax, and snakebites treatment in 1500 ml of hexane. The contents were kept for 3 days away from direct sunlight, undisturbed, then filtered through sterile filter paper. (Kabir et al., 2005). Different communities in Africa use The filtrate was concentrated at 38.5 to 42°C. parts of Carissa edulis to alleviate pain, treat venereal diseases, glandular inflammation, induce abortion and restore virility (Githiori et al., 2004). In Kenya and Preparation of ethyl acetate extract Tanzania decoction from Launae cornuta roots is used as The hexane residues were re-soaked in 1500 ml of ethyl acetate. a remedy for cough, typhus fever and measles The contents were kept for 5 days away from direct sunlight, (Schippers, 2004). undisturbed and afterward filtered. The filtrate was concentrated at This study investigated the anti-Salmonella activities of 38.5 to 42°C. T. diversifolia, W. ugandensis, C. megalocarpus, C. edulis and Launae cornuta. Clinical isolates of S.ser.Typhi (ATCC 13347), S.ser.Typhi (ATCC 43579), Preparation of methanol extract S.enterica (ATCC 2162) and S. ser. Typhimurium (ATCC The ethyl acetate residues were re-soaked in 1500 ml of methanol 1408) were used in the study. and kept for 36 h away from direct sunlight undisturbed. After filtration, the filtrate was concentrated at 65°C. The extracts were stored at 4°C until used. MATERIALS AND METHODS Salmonella strains Preparation of aqueous extract Clinical samples of S.ser.Typhi (ATCC 13347), S.ser.Typhi (ATCC Five hundred grams (500 g) of each of the powdered plant 43579), S.enterica (ATCC 2162) and S. ser. Typhimurium (ATCC materials was weighed and soaked separately in 1500 ml of 1408) were provided by the Centre of Microbiology Research, distilled water. The contents were warmed in a water bath for 2 h at 256 J. Med. Plants Res. Table 1. Profile of the five medicinal plants. Botanical name Family name Part of the plant used Tithornia diversifolia Solanaceae Flowers and leaves Warburgia ugandesis Conellaceae Roots and stem barks Croton megalocarpus Euphorbiaceae Barks Carissa edulis Apocynaceae Roots and barks Launae cornuta Asteraceae Roots and leaves 60°C, then left to stand at room temperature for 10 h, undisturbed. Statistical analysis They were subsequently sterile filtered and filtrate freeze dried to powder. The powder were weighed and stored until used. Anti-Salmonella activity was determined from means of triplicates in zones of inhibition and duplicates in MICs. Collected data was analysed statistically using one way ANOVA (SAS, Version 9.0). Determination of phytochemical constituents Difference in values at P < 0.0001 were considered statistically significant. The freshly prepared extracts were subjected to standard phytochemical analyses for tannins, alkaloids, terpenoids, flavanoids, glycosides, steroids and saponin as described by Jigna RESULTS et al. (2006). Out of 36 plant extracts screened using disc diffusion Controls assay, only nine extracts inhibited
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