Applied Biology and Biochemistry Publications

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Quantitative Amino Acid Analysis of Extracted Soyabean Meal Hydrolysates by Ion Exchange Chromatography
(Zimbabwe agriculture Journal., 1985) Siwela, Andrew H.
Although there are tables of amino acids In foods and feeding stuffs written by authorities in the field, it is felt that there is a need for such tables appertaining to Zimbabwean conditions. As a first step to this objective a survey was undertaken to determine the amino acid cr;mtent, and its variability, of extracted soyabean meal which in this country is used as a majO'/' source of protein in feedstuffs in common with worldwide practice. The variability from batch to batch Wlm' found. t,o be very low.
The effect of schistosomiasis on the activation of aflatoxin B1
(1986-03) Hasler, Julia A.; Siwela, Andrew H.; Nyathi, C.B.; Chetsanga, C.J.
This study examined activation of aflatoxin Bl (AFB1) in livers of Schistosomamansoni-infected and noninfected mice by measuring covalent binding of ('H]AFBl to cellular macromolecules in vivo and in vitro. During a one week time period after AFB1 treatment of animals, maximal binding of ('H]AFB1 to DNA, RNA and protein in liver occured during the 1 - 6 hour period after treatment, with less binding throughout of AFB1 to macromolecules of infected mice. Experiments performed in vitro to determine the capacity of liver microsomes to mediate the binding of AFB1 to calf thymus DNA showed that microsomes from infected mice mediated the binding of less ('H]AFB1 to DNA than those from noninfected animals.
Aflatoxin Contamination of Stored Groundnuts in Zimbabwe.
(ICRISA T (International Crops Research Institute for the Semi-Arid Tropics)., 1989) Siwela, Andrew H.; Caley, A.D.
Aflatoxins were analyzed in groundnuts stored for either local or export sales in the period 1982/83 to 1986/87. Four hundred and forty-one samples of seven groundnut varieties were collected for analysis. Sixty-eight percent of the samples had total concentrations of aIlatoxin BJ and OJ of up to 25 JIg kg-J. In the 1986/87 groundnut crop, the Flamingo and Makulu Red varieties, which constitute the bulk of export sales, had up to 25 JIgkg-J aIlatoxin BJin 79%of the samples, plus aIlatoxin OJ in 67% of the samples. Overall, the Egret variety was the most susceptible to aIlatoxin contamination during this period.
The effect of schistosomiasis on the covalent binding of 2-acetylaminofluorene to mouse liver macromolecules in vivo and in vitro
(Elsevier, 1990-07-15) Siwela, Andrew H.; Nyathi, C.B.; Chetsanga, C.J.; Hasler, Julia A.
The covalent binding of [14C]acetylaminofluorene (AAF) to macromolecules in vivo and in vitro was measured in Schistosoma mansoni-infected and in non-infected mice. Liver microsomes from infected mice demonstrated a 42% decreased capacity to mediate covalent binding of AAF to DNA. In addition, the extent of binding of AAF to liver macromolecules in vivo was generally less in infected than non-infected mice.
Effects of phenobarbital and 3-methylcholanthrene pretreatment on the pharmacokinetics of praziquantel in rats.
(1992-06-25) Masimirembwa, Collen M.; Naik, Yogeshkumar S.; Hasler, Julia A.
The effects of phenobarbital (PB) and 3-methylcholanthrene (MC) pretreatment on the phannac~kinetics of praziquantel (PZQ), a schistosomicide were studied in Spnrgue-Dawley rats. Blood samples at different time intervals were obtained by severing the tail vein and were analyzed for unchanged PZQ by HPLC. The PB-pretreated rats showed a &fold decrtase in AUC, a 5-fold decrease in Cmax and an 8-fold increase in Ch compared to the saline treated controls. The MC-pretreated rats and their oliveoil treated controls did not show any statistically significant diffmnces in the above parameters. These results suggest that PZQ is extensively metabolised by PB-inducible cytochrome P-450 isoforms and not by MC-inducible isoforms. These findings also suggest that the bioavailability of praziquantel could be altered to a significant extent in humans taking drugs that are phenobarbital type inducers.
Xenobiotic metabolizing enzymes of the freshwater snails.Planorbella and lymnaea (RADIX)Natalensis.
(J. Med. and Appl. Malacol, 1994) Naik, Yogeshkumar S.; Magwere, Tapiwanashe; Matiyenga, M.; Hasler, Julia A.
Very little is known about the ability of snails to metabolise and remove xenobiotics such as molluscicides. The present study was conducted to determine whether the freshwater snails Planorbella duryi and Lymnaea (Radix) nntaIensis possess two of the major enzyme systems of detoxication, namely the P450 monooxygenase system and the glutathione Stransferases. We were able to measure microsomal cytochrome b5 and NADPH cytochrome c reductase activity, indicating a functional mixed function oxidase system. However, probing with three known substrates for the mammalian enzyme, we were unable to detect any cytochrome P450 mediated activity. Glutathione Stransferase activity was detectable in cytosolic fractions. In general, the activities detectable in snails were much lower than in mammalian liver preparations. The existence of these enzymes in snails suggests that studies should be undertaken to observe the interaction between these enzymes and xenobiotics such as candidate molluscicides. Key words: Planorbella duyi, Lyrnnaea (Radix) natalensis, cytochrome P450, glutathione
Combined use of phenyl-bonded phase clean-up and HPLC for the determination of aflatoxins
(Tropical Science, 1996) Siwela, Andrew H.
Phenyl·bonded phase clean-up and HPLC were used to measure aflatoxins in groundnuts, peanut butter, beans, cowpeas and maize meant for human consumption. Of 277 samples, 84% were al or bleow the 20 pp.b. limil. Of 100 animal feed samples, 28% were at or below 5 p.p.b., the proposed limit infeedfor milk-producing animals.
A Comparison Of Organochlorine Pesticide Residues In Upper Ncema And Lower Umguza Dams, Zimbabwe.
(Journal of Applied Science in Southern Africa, 1996) Siwela, Andrew H.; Marufu, G.; Mhlanga, A.T.
This study examined the levels of chlorinated hydrocarbons (organochloride pesticides) in the ecosystem of Ncema and Umguza dams. Samples of water, water weeds, mud and fish were collected from each dam. The fish species collected from the two dams were river sardine (Mesobo/a brevianus), branded bream (Tilapia sparmanii), African catfish (e/arias gariepinus) and red-breasted bream (Tilapia rendelli). The samples were analysed for hexachlorocyclohexane, DDT, and its metabolites, DOE and DOD, aldrin and dieldrin. DDT was detected in all the samples, its concentration ranging from 1,12 J.l9 per kg in water weeds to 265 J.l9 per kg in mud from Upper Ncema dam. Aldrin was also detected in all the samples (except for branded bream from Upper Ncema dam), the highest level __being 223 ~g per kg in_ ~atdln~$'-rom Lower Umguza dam. Traces.of hexachlorocyclohexane were detected in some of the samples, the highest level of 20,1 ~g per kg in African catfish from Upper Ncema dam. Dieldrin was detected only in mud samples from Lower Umguza dam at a concentration of 80,5 ~g per kg. Overall, this study indicates that Upper Ncema dam has a higher pesticide load compared to Lower Umguza dam.
Effects of chloroquine treatment on antioxidant enzymes in rat liver and kidney.
(Elsevier Inc., 1996-05-09) Magwere, Tapiwanashe; Naik, Yogeshkumar S.; Hasler, Julia A.
The effect of chloroquine (CHQ) administration on antioxidant enzymes in rat liver and kidney was studied. Male Sprague-Dawley rats were administered 20 mglkg CHQ once a week for 4 weeks (chronic treatment) or a single dose at 10 or 20 mglkg (acute treatment). Antioxidant enzyme activities were determined in cytosolic fractions of liver and kidney, whereas reduced glutathione (GSH) and malondialdehyde (MDA) were determined in tissue samples. Results indicate minimal effects of acute CHQ treatment, whereas chronic treatment with CHQ differentially affected antioxidant enzymes in the two organs. Superoxide dismutase activity was increased nearly twofold, while activities of selenium glutathione peroxidase (GPX), catalase, and NAD (P) H: quinone oxidoreductase were decreased in livers of CHQ-treated rats compared to controls. No significant effects of CHQ on glutathione reductase, GSH, and MDA levels were seen in the liver. Fewer effects of CHQ were observed in the kidney where a decrease in GPX activity and an increase in MDA levels was seen. Lowering of antioxidant enzymes activities in the liver by CHQ could render the organ more susceptible to subsequent oxidative stress; while increased MDA production after CHQ treatment in the kidney indicate that the organ is being subjected to oxidative stress. This could have implications for prolonged chloroquine intake.
Phenotyping of the glutathione S-transferase M1 polymorphism in Zimbabweans and the effects of chloroquine on blood glutathione S-transferases M1-A
(Elsevier, 1997-05-31) Mukanganyama, Stanley; Masimirembwa, Collen M.; Naik, Yogeshkumar S.; Hasler, Julia A.
The frequency of the null allele phenotype of glutathione S-transferase (GST) M1 was investigated in 114 Zimbabweans and results for a subset of 63 subjects were compared with genotyping by PCR. In addition, the effect of the antimalarial chloroquine on blood levels of GSTMl and GSTA in 19 subjects was studied. Quantification of GSTs was by enzyme linked immunosorbent assays (ELISA). Thirty percent of the subjects were of the GSTMl null phenotype. Comparison of results of phenotyping by ELISA and genotyping by PCR showed that 16% of samples were in discordance; unknown mutations in the GSTMl gene in the Zimbabwean population may explain this observation. Chloroquine decreased levels of blood GSTM1 and GSTA by 50% or more. In populations treated with chloroquine, these decreases in GST activities might lead to compromised ability to detoxify xenobiotics, could confound GSTMl phenotyping and might invalidate use of GSTA as an indicator of liver damage. O 1997 Elsevier Science B.V. Keywords: Glutathione S-transferases; Phenotype; Black Zimbabweans; Chloroquine
Effect of schistosome infection on hepatic drug metabolising enzymes
(South African Journal of Science, 1998-06) Naik, Yogeshkumar S.; Hasler, Julia A.
Schistosomiasis is a parasitic disease that affects over 200 million persons worldwide.' The disease is caused by infection with any one of several species that are known to affect humans. Infected persons are likely to be consuming a wide spectrum of xenobiotics such as drugs and environmental toxins. The drugs consumed would not only include praziquantel and other schistosomicides, but also those used for the treatment of other parasitic diseases such as antimalarials, anthelmintics and antibiotics. Experiments involving humans and experimental animals suggest that infection with schistosomes causes a reduction in the host's ability to metabolise and remove drugs from the body.
Effect of schistosome infection on hepatic drug metabolising enzymes
(1998-06) Naik, Yogeshkumar S.; Hasler, Julia A.
Schistosomiasis is a parasitic disease that affects over 200 million persons worldwide.' The disease is caused by infection with any one of several species that are known to affect humans. Infected persons are likely to be consuming a wide spectrum of xenobiotics such as drugs and environmental toxins. The drugs consumed would not only include praziquantel and other schistosomicides, but also those used for the treatment of other parasitic diseases such as antimalarials, anthelmintics and antibiotics. Experiments involving humans and experimental animals suggest that infection with schistosomes causes a reduction in the host's ability to metabolise and remove drugs from the b~dy.~.~ The metabolic fate of drugs is dependent to a large extent on the expression and activity of the microsomal drug metabolising enzyme^.^^^ These enzymes include the microsomal cytochrome P-450 dependent monooxygenase system and the uridine diphosphate glucuronosyl transferases as well as other cytosolic enzymes such as glutathione S-transferases. This article reviews the effects of experimental Schistosoma mansoni infection on hepatic drug metabolising enzyme activity.
Regulatory aspects of aflatoxin control in Zimbabwe - a review
(JASSA, 1999) Siwela, Andrew H.; Nziramasanga, Nozipho
Aflatoxins are highly toxic and carcinogenic chemicals produced by fungi of the Aspergillus family. As the fungi may be found growing on virtually all stored agricultural commodities and foodstuffs, many countries have put in place regulations to control the levels of afJatoxins and other mycotoxins that may be found contaminating foodstuffs and stock feeds. Current regulations and aflatoxin control in Zimbabwe is based on a voluntary code of conduct drawn in 1971 which putthe action limit at between 4f,glkg to 20!.glkg aflatoxin G, (AFG,J or Sf,glkg to 2Sf,glk9 aflatoxin B, (AFB1) for human consumption. Any commodity contaminated by Aflatoxin greater than these limits should be used for stock feed and those below these limits could be used for the export market. The 2Sfl9/kg AFB, action limit has been levi sed to 20f'9/kg by the statutory instrument of 1990 under the Food Standard Act of 1971. This paper reviews work that has been done in the past ten years in the monitoring and control of aflatoxin in foodstuffs and feeds. Aflatoxin levels in foods, stock feeds, urine and human milk from a cross section of the population have shown that the population and livestock are exposed to aflatoxin. Disease outbreaks in livestock have also been linked to aflatoxins and other mycotoxins. It is recommended that aflatoxin monitoring be intensified in support of the Food Standards Act and the voluntary code of conduct be made into a legal instrument to control transmission of aflatoxin in the food chain.
Chemical composition and physical characteristics of cowpea haulms as forage for ruminants
(South African Society of Animal Science, 2000) Mokoboki, H.K.; Ayisi, K.K.; Ndlovu, Lindela R
Cowpea ( Vigna unguiculata) is a major grain legume grown by small- scale farmers in the Northern Province of South Africa. The crop is indigenous to Africa and is used extensively in many cropping systems throughout Africa (Eaglesham et al., 1981). Cowpea straws are potential supplementary feed for ruminants in the dry season. Cowpea straws are likely to have higher crude protein than cereal straws and may thus improve digestibility of cereal straws if used together. However, cowpea straws may contain anti-nutritional factors such as tannins and total phenols which affect animal performance. The protein and tannin content of cowpea straws is likely to be affected by variety and growth environment. Straws of varieties adapted to Northern Province have not been evaluated for their content of these constituents. Whilst animal intake and utilisation are the ultimate test of forage suitability, the procedures are unsuitable when large number of forage germ-plasm are to be screened. Chemical and physical characteristics that are known to affect intake and utilisation are often used in such circumstances. The main aim of this research was to determine the effect of variety on physical and chemical characteristics of straws of long season cowpeas and also to determine if haulms of varieties that retain greenness longer will have a better nutritional value than those that dry early.
Establishment of baseline values of copper, zinc and manganese in ostriches in Zimbabwe: a preliminary study.
(Zimbabwe Vet Journal, 2000) Siwela, Andrew H.; Shonhai, A.; Foggin, C.M.; Revol, B.
Ostrich (Struthio came/us) sera from apparently 302 healthy birds on 32 different fanns around the country were analysed for three essential trace elements (copper, zinc and manganese) by atomic absorption spectrometry with the objective establishing baseline values which could be used as guidelines in the diagnosis of deficiency or toxicity of the trace elements in ostriches. Trace elements levels in ostrich sera were also detennined according to age groups i.e chicks, growers and b~eeders. Comparisons were also done to determine whether the geographical regions had any influence on the levels of the trace elements in the different categories of birds. Results indicated significant variations with regard to age and geographical region variations in the level of copper. No significant age variations were evident for manganese levels but regional differences were noted. Zinc levels did not show significant geographical differences but variation in the concentration of the trace element according to age was strongly evident.
Proposed Reductive Metabolism of Artemisinin by Glutathione Transferases in vitro
(The Harwood academic publishers imprint,, 2001-01-18) Naik, Yogeshkumar S.; Muganyama, Stanley; Hasler, Julia A.; Widersten, Mikael; Mannervik, B.
Artemisinin is a sesquiterpene lactone containing an endoperoxide bridge. It is a promising new antimalarial and is particularly useful against the drug resistant strains of Plasmodium fakiparum. It has unique antimalarial properties since it acts through the generation of free radicals that alkylate parasite proteins. Since the antimalarial action of the drug is antagonised by glutathionc and ascorbate and has unusual pharmacokinetic properties in humans, we have investigated if the drug is broken down by a typical reductive reaction in the presence of glutathione transferases. Cytosolic glutathione transferases (GSTs) detoxlfy electrophilic xenobiotics by catalysing the formation of glutathione (GSH) conjugates and exhibit glutathione peroxidase activity towards hydroperoxides. Arternisinin was incubated with glutathione, NADPH and glutathione reductase and G~TiSn a coupled assay syste&analogous to the standard assav scheme with cumene hv,d rover- L oxide as a substrate df GSTS. Arternisinin was shown to stimulate NADPH oxidation in cytosols from rat liver, kidney, intestines and in affinity purified preparations of GSTs from rat liver. Using human recombinant GSTs hetelorogously expressed in Escherichia coli, artemisinin was similarly shown to stimulate NADPH oxidation with the highest activity observed with GST MI-1. Using recombinant GSTs the activity of GSTs with artemisinin was at least two fold higher than the reaction with CDNB. Considering these results, it is possible that GSTs may contribute to the metabolism of artemisinin in the presence of NADPH and GSSG-reductase We propose a model, based on the known reactions of GSTs and sesquiterpenes, in which (1) artemisinin reacts with GSH resulting in oxidised glutathione; (.2) the oxidised glutathione is then converted to reduced glutathione via glutathione reductase; and (3) the latter reaction may then result in the depletion of NADPH via GSSG-reductase. The ability of artemisinin to react with GSH in the presence of GST may be responsible for the NADPH utilisation observed in vitro and suggests that cytosolic GSTs are likely to be contributing to metabolism of artemisinin and related drugs in vivo
Inhibition of Glutathione S-Transferases by antimalarial drugs possible implications for circumventing anticancer drug resistance.
(Wiley-Liss, Inc., 2001-08-06) Mukanganyama, Stanley; Widersten, Mikael; Naik, Yogeshkumar S.; Mannervik, B.; Hasler, Julia A.
A strategy to overcome multidrug resistance in cancer cells involves treatment with a combination of the antineoplastic agent and a chemomodulator that inhibits the activity of the resistance-causing protein. The aim of our study was to investigate the effects of antimalarial drugs on human recombinant glutathione S-transferase (GSTs) activity in the context of searching for effective and clinically acceptable inhibitors of these enzymes. Human recombinant GSTs heterologously expressed in Escherichia coli were used for inhibition studies. GST Al-l activity was inhibited by artemisinin with an IC,, of 6 pM, whilst GST MI-l was inhibited by quinidine and its diastereoisomer quinine with IC5,s of I2 pM and 17 pM, respectively. GST M3-3 was inhibited by tetracycline only with an IC,, of 47 pM. GST PI-l was the most susceptible enzyme to inhibition by antimalarials with IC,, values of I, 2, 1, 4, and 13 pM for pyrimethamine, arteniislnin, quinidine, quinine and tetracycline, respectively. The IC,, values obtained for artemisinin, quinine, quinidine and tetracycline are below peak plasma concentrations obtained during therapy of malaria with these drugs. It seems likely, therefore, that GSTs may be inhibited in vivo at doses normally used in clinical practice. Using the substrate ethacrynic acid, a diuretic drug also used as a modulator to overcome drug resistance in tumour cells, GST PI-l activity was inhibited by tetracycline, quinine, pyrimethamine and quinidine with IC,, values of 18, 27, 45 and 70 pM, respectively. The ubiquitous expression of GSTs in different malignancies suggests that the addition of nontoxic reversing agents such as antimalarials could enhance the efficacy of a variety of alkylating agents.
Prevalence of Paramphistomes in Mashonaland West, Central, and East, and Midlands Provinces, Zimbabwe
(Acta ZoologicaTaiwanica, 2002) Dube, S.; Siwela, Andrew H.; Masanganise, K.E.; Dube, C.
Paramphistomes were collected from cattle from various localities in Mashonaland West, Central, and East and Midlands Provinces. After slaughter, their rumens and reticulum were cut open and examined for paramphistomes. Identifications were based on the morphological structuresand measurementsof diagnosticfeaturesfrom flattened and median sagittal sections. Analysis of the structures of the acetabulum, pharynx, and genital atrium revealed the following parasites in the area, Paramphistomum microbothrium, P. clavula, Calicophoron calicophorum, C. raja, and Gigantocotyle symmeri. The average number of cattle found infected annually was 29.39% with an upper limit of the range of around 69%. Parasite loads and damage to tissues were not serious in most cases, except in a few severe cases where the papillae were damaged and a catarrhal pus-like exudate was noticed on the mucosa
Molecular characterisation of Armillaria species from Zimbabwe
(The British Mycological Society, 2003-01) Mwenje, Eddie; Wingfield, Brenda D.; Coetzee, Martin P.A.
Armillaria species are amongst the most important pathogens of trees and have a world-wide distribution. In recent years, the taxonomy of Northern Hemisphere Armillaria spp. has been extensively treated, but those occurring in Africa are poorly known. Previously, isolates of Armillaria from Zimbabwe have been grouped based on morphology and biochemical tests. In this study, six isolates representing the three previously characterized groups of Armillaria spp. occurring in Zimbabwe were analysed using DNA-based techniques. Three distinct clusters emerged from both PCR–RFLP and analysis of sequence data for the IGS-1 rRNA operon. The three groups corresponded to those previously identified based on morphology and biochemical tests. Differences in IGS-1 sequences strongly suggest that the Zimbabwean groups represent three distinct taxa. Isolates belonging to Group I, previously assumed to be to A. heimii, were similar to those identified as A. fuscipes from South Africa and La Reunion. Group II isolates resided in a clade apart from all other isolates and appear to represent A. heimii. The remaining isolates residing in Group III clustered with isolates from Zambia and Cameroon. These are different from A. heimii and A. fuscipes and apparently represent an undescribed taxon.
Decontamination of aflatoxin-contaminated maize by dehulling
(Society of Chemical Industry., 2005) Siwela, Andrew H.; Siwela, Mthulisi; Matindi, Gibson; Dube, S.; Nziramasanga, Nozipho
Dehulling of maize grains as an aflatoxin decontamination method was investigated. Sixty kilograms of maize (whose average moisture content was 110 g kg−1) were thoroughly mixed and divided into two samples. The kernel moisture content of one sample was adjusted to 200 g kg−1 while the other (control) was left at 110 g kg−1. The two samples were kept at ambient temperature (25–30 ◦C) for 21 days. Twenty sub-samples, ten from each, were further divided into two so that one lot was dehulled while the other was not dehulled. These sub-samples were then milled to pass through a 1mm screen and the meal was evaluated for aflatoxin contamination. It was found that there was a 92% decrease in aflatoxin levels in dehulled maize meal compared with undehulled maize meal. We therefore suggest that dehulling the grain can be used to reduce aflatoxin levels in maize.

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