Masters Degrees (Physiology)

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The effects of nerve stimulation on pacemaking activities of biological tissues.
(1973) Bhagat, Chotoo Ichharam.; Reid, John Victor Oswald.
The effects on the cardiac cycle length of stimulating the vagus nerves with single supramaximal electrical shocks depended upon when they were stimulated during the cycle. A maximum prolongation of the cardiac cycle was obtained when the vagi were stimulated 167 msec (SD±64) after the peak of an electrocardiogram P wave. The interval between a P wave and the subsequent vagal stimulation was called Pl-St interval. Pl-St(max) was the Pl-St interval at which maximum prolongation of the cardiac cycle occurred. Pl-St(max) increased significantly (p (0.001) with longer cardiac cycles. When the Pl-St intervals were shorter or longer than 167 msec (SD±64) the effects of vagal stimulation were less. The latent period for the effects of vagal stimulation was 195 msec (SD±32) The latent period also increased significantly (p(O.Ol) with longer cardiac cycles. The rise time of the vagal effect, obtained by subtracting (Pl-St(max)+ latent period) from the control cardiac cycle length, was 124 msec (SD+31) and occurred between Pl-St intervals of 167 msec (SD±64) and 291 msec (SD±70). The rise time did not vary with cardiac cycle length (p) 0.1), but the magnitude of the maximum response to vagal stimulation was inversely proportional to rise time (p <. 0.02). The peak response to vagal stimulation must have occurred when the vagal effects pegan somewhere in the middle of diastolic depolarization of the pacemaker cells in the S-A node. The reasons for this were discussed. The half-decay time for the effects of vagal stimulation was 210 msec (SD±102). The slope of the curve relating the prolongation of the cardiac cycle length to Pl-St is positive at Pl-St intervals less than 167 msec (SD±64) and negative at Pl-St intervals between 167 msec (SD±64) and 291 msec (SD±90). The positive slope ranged from 0.13 to 0.48 with a mean of 0.23. The paradoxical responses of the S-A node to vagal inhibitory input obtained by Reid (1969), Levy et al (1969)and Dong and Reitz (1970) would be explained by the dependence of the cardiac cycle length upon the time of arrival of vagal stimulus in relation to the previous P wave and upon the slope of the curve relating the prolongation of the cardiac cycle length to Pl-St interval being positive and between zero and two at Pl-St intervals less than 167 msec (SD±64. The effects of single shock stimulation of the vagus nerves persisted for 3.890 sec (SD+l.255)7 the number of cardiac cycles involved varied between 5 and 11. The duration of the effects of vagal stimulation did not depend upon when during the cardiac cycle the vagi were stimulated. A "dip" in the response to vagal stimulation was present in all the experiments. The possibility of the "dip" phenomenon being due to simultaneous stimulation of the sympathetic fibres in the vago-sympathetic trunk was ruled out. It is suggested that the "dip" phenomenon may be due to transient accumulation of K+ in the interstitial fluid surrounding the pacemaker cells in the S-A node.There was no paradoxical response of the smooth muscle in the distal colon of the adult rabbit when the frequency of sympathetic inhibitory input was continuously increased. A paradoxical response in the frequency but not in the size of the contraction of the smooth muscle was obtained when the sympathetic nerves were stimulated with bursts of stimuli, each burst consisting of 5-40 impulses, 10 msec apart. One may conclude from this that the delay of the next spontaneous contraction but not the inhibition of the size of smooth muscle contraction is dependent upon the arrival time of a burst of stimuli during a contraction cycle. This was confirmed in an experiment when the sympathetic nerves were stimulated with single bursts of stimuli applied at different times during the contraction cycle. It is unlikely that such a paradoxical response would occur under physiological conditions as this would require the natural sympathetic efferent discharges to the smooth muscle to occur in regular bursts, each burst consisting of impulses at a high frequency. Stimulation of the sympathetic nerves at 3, 5, 10 and 25 PPS caused an inhibition of the size and frequency of smooth muscle contraction in the distal colon of the newborn rabbit. Assuming that the cholinergic fibres are excitatory there is therefore no evidence for the sympathetic fibres to the distal colon being cholinergic in the newborn rabbit. This is contrary to Burn's (1968) report of the sympathetic fibres being motor and cholinergic to the small intestinal smooth muscle in the newborn rabbit.The heart rate increased rapidly at the onset of exercise and then more gradually over the rest of the exercise period. The initial increase in the heart rate during exercise was not affected by adrenergic blockade but the subsequent increase in heart rate was significantly reduced by adrenergic blockade. Hence the increase in heart rate at the onset of exercise is due primarily to a decrease in the cardiac vagal efferent discharge, whereas the subsequent increase in heart rate is due to both a further decrease ln vagal discharge and an increase in sympathetic discharge to the S-A node. In almost all the sub jects there was initially a rapid decline in the heart rate in the post-exercise period, but subsequently the heart rate returned to resting levels in a variety of ways. These were classified into 5 types. Of particular interest to the present study was the Type V pattern of heart rate change. This was characterised by an increase in heart rate of 6 beats or more per minute during the post-exercise period, with or without superimposed arrhythmia. The Type V pattern may be the equivalent of the paradoxical responses to inhibitory input demonstrated in animal experiments i.e. an increase in the heart rate with increasing vagal stimulation frequency. Type V pattern occurred more frequently at mild exercise levels (4 out of 14) than at moderate exercise level (lout of 14) and also more frequently in adrenergic blocked individuals (11 out of 28) than in control subjects (5 out of 28) It is suggested that the sympathetic effects on the P-R interval and arterial baroreceptor modulation of vagal efferent discharge protect again st the occurrence of paradoxical responses to vagal inhibitory input. They may do so by confining the vagal discharge to the rise time of vagal effect during the cardiac cycle. On the other hand the Type V pattern in p-adrenergic blocked individuals may be due to a decrease in the vagal discharge, in which case Type V pattern would not be a paradoxical response. The changes in minute ventilation in the post-exercise period were also variable. Besides a gradual decline in minute ventilation there were also gradual increases and sudden increases and decreases in minute ventilation. These may represent a form of paradoxical response to increasing inhibitory input and decreasing excitatory input to the respiratory neurones in man. However, all the changes in minute ventilation could also be explained by fluctuating excitatory and inhibitory neural input to the respiratory neurones.
A quantification of heat load as assessed by indicators of tissue damage in rats.
(1984) Manjoo, Mahomed.; Burger, F. J.; Kielblock, Artz Johan.
Heatstroke is an illness that occurs when body temperature is grossly elevated, causing widespread tissue damage. The extent of tissue damage depends on the level of body temperature elevation and the duration. Despite the fact that the diagnosis of heatstroke is based on sound scientific principles, namely the elevation of serum enzyme levels as indicators of tissue damage, the sensitivity of these parameters of tissue damage in the prodromal period of heatstroke is less well established, especially for sub-lethal stress conditions. Furthermore, it is not known to what extent given elevations in serum enzyme levels reflect the nature of various combinations of hyperthermia and its duration as sustained during the prodromal period. In an attempt to throw some light on the questions posed above anaesthetized rats were exposed to three different sets of thermal conditions. However, the amount of heat gained over and above baseline levels was controlled to a 20% rise irrespective of the experimental conditions. Above this increment animals did not survive thus indicating excessive stress. Plasma enzyme levels were assayed in each group of animals upon termination of stress, six hours post-stress and 24 hours post-stress in order to investigate the patterns of enzyme release as well as the sensitivity of the respective indicators of tissue damage. On the basis of plasma enzyme assays, the tissue damage sustained during these particular experimental conditions was mild to moderate, completely reversible, not indicative of heatstroke but merely of generalized tissue damage. The results suggest that in addition to the established positive relationship between the level and duration of hyperthermia and tissue damage, a third component,namely the rate of rise in body temperature, nay constitute an important factor in the ultirrate pathology. In this regard, i.e. sub-lethal stress, creatine kinase proved to be the most sensitive and, therefore, the most useful parameter of tissue damage.
The effect of vitamin B-6 deficiency on copper, zinc and iron balance in the rat.
(1988) Channa, Mahendra Lala.; Burger, F. J.
In order to address the contradictory reports on the rat trace element status during a vitamin 8-6 deficiency, Zn, Cu and Fe balance was assessed over 8 weeks in 3 groups of young male rats. Group 1 was the control group fed on a diet supplemented with 3,5 mg/kg of vitamin B-6. Group 2 was the marginally deficient group, fed a diet supplemented with 1,75 mg/kg of vitamin B-6 and Group 3 was the totally deficient group without any vitamin 8-6 in the diet. Diet, urine and fecal samples were analysed to determine the mean daily Zn, Cu and Fe balance for each group during each week of the study. Urinary xanthurenic acid (XA), plasma pyridoxal (PL), and plasma pyridoxal 5' -phosphate (PLP) were also analysed in order to determine the vitamin 8-6 nutritional status of each animal group. The totally deficient Group 3 showed decreased growth and food consumption when compared to the control and marginally deficient groups. There was a significant difference in XA excretion and plasma PLP and PL levels between all 3 groups with a progressive increase in XA excretion and a progressive decrease in PLP and PL levels from Group to Group 3. Although all groups were found to be in a positive balance for Zn, Cu and Fe, the totally deficient group was in a less positive balance compared to Groups 1 and 2. Furthermore, the percentage excretion of Zn and Cu, when compared to the intake, was greater in this group. This increased excretion, coupled with a decreased food intake, accounted for the lowered Zn and Cu balances in the totally deficient group. Fe excretion did not differ significantly between all 3 groups. The marginally deficient Group 2 showed no significant difference in growth, food consumption, trace element balance and excretion when compared to the control group, indicating the beneficial effects of even subminimal levels of vitamin B-6. This study confirms that the nutritional status of t~ace elements, especially that of Zn and CU, is related to the vitamin B-6 status of the animal.
The effect of vitamin B-6 deficiency on the levels of tissue copper, iron and zinc in the rat.
(1989) Mackraj, Irene.; Burger, F. J.
No abstract available.
The effect of dietary egg on human plasma cholesterol and triglyceride levels.
(1990) Raidoo, Kogie.; Burger, F. J.
No abstract available.
The effect of vitamin B-6 deficiency on the bioavailability of zinc in the rat.
(1990) Moodley, Dhanabaikum.; Burger, F. J.
No abstract available.
Studies on the isolation of the polymerase genes from the H1N1 influenza A virus.
(1992) Naidoo, Richard.; Manning, D.
Vaccines directed against the influenza virus become ineffective due to continuous mutation. An alternative approach might be to control replication at the genomic level by enzymatic methylation of the polymerase genes. Hence in this study, a method to locate and successfully isolate the H1N1 influenza A polymerase genes was investigated. The virus was cultured in chick embryos via the allantoic route using aseptic techniques. Following incubation, the allantoic fluid was isolated and washed to remove any contaminating blood cells. The allantoic fluid was checked for fungal and bacterial contamination using the blood agar test and the presence of the virus was established by the haemagglutination titration test. Viral particles were pelleted by ultracentrifugation. Electron microscopy verified the morphology and size of these viruses while immunofluorescence studies, using a monoclonal antibody, confirmed the influenza A strain. The ribose test verified the presence of RNA in the samples. Purified viral pellets were pooled and homogenised in buffer containing guanidine thiocyanate, mercaptoethanol and sarkosyl. The samples were incubated on ice before mechanical disruption of the virus. Viral RNA was isolated from the upper aqueous layer after a standard phenol/chloroform extraction procedure. RNA was quantified spectrophotometrically and purity assessed initially by the absorbance ratio readings at 260/280 nm. Electrophoresis of the RNA samples was performed together with RNA molecular weight markers on a 1.5% formamide agarose gel. Five bands were identified and the band containing the polymerase genes was size selected, located and excised. Purification of the polymerase genes from the agarose was achieved by using the BIO 101 RNAid kit. The three isolated polymerase RNAs were reverse transcribed using the Boehringer Mannheim cDNA synthesis kit. The results indicate that the H1N1 influenza virus was successfully grown and isolated from chick embryos. Absence of contamination and verification of viral presence at different stages of the study were indications that asepsis was successfully achieved. The RNA obtained was sufficient and suitable for cDNA synthesis. This cDNA may now be used for further molecular analysis and subsequent DNA methylation studies. Further, transfection studies may then be performed to determine, if any, the the expression of methylated and unmethylated cDNA.
The interrelationship of dietary cholesterol, copper and zinc on plasma lipids and tissue copper and zinc levels in the rat.
(1992) Nadar, Anand.; Burger, F. J.
No abstract available.
The interrelationship of dietary cholesterol, copper and zinc on plasma lipids and tissue copper and zinc levels in the rat.
(1992) Anand, Nadar.; Burger, F. J.
No abstract available.
The Use of an enzyme-linked immunosorbent assay (Elisa) for the determination and characterization of antiendotoxin antibodies.
(1994) Badsha, Nasima.; Gaffin, Stephen L.
Recent clinical studies have highlighted the effectiveness of immunotherapy for Gram-negative bacteraemia in humans. Studies in America, undertaken on patients with Gram-negative bacteraemia, have shown that mortality was reduced by virtually 50% in patients who received specific antiendotoxin antiserum. In India, mortality from pseudomonas septicaemia was significantly reduced by the administration of small quantities of a anti-pseudomonas immunoglobulin. The antibodies in those studies were raised by vaccination of healthy volunteers with heat-killed Gram-negative bacteria or vaccines containing endotoxin. Adverse side effects in volunteers as well as logistic and legal problems make it difficult to produce antiserum on a large scale, in this manner. In Israel, S.L. Gaffin and coworkers found that approximately 7% of plasma units in a blood bank had antiendotoxin antibody concentrations of 40 ).1g/m1 or greater. This high titre human plasma significantly protected cats from lethal endotoxic shock secondary to haemorrhage. The immunoprecipitin technique used by them to measure antiendotoxin antibody concentrations was unsuitable for screening large numbers of blood samples. To overcome this problem we have devised an enzyme-linked imounosorbent assay (ELISA) for determining the level of antiendotoxin immunoglobulin G in human plasma. The assay, which is suitable for large scale use, was found to be specific for antiendotoxin antibodies. It was calibrated using a serum sample of specific antibody concentration as determined by an ilununoprecipitin assay. Serum samples found to be high in antiendotoxin titres (> 40_ug/m1) were tested for their specificity towards endotoxins from 12 bacterial iv strains and species. While each sample was found to have its own characteristic specificities, most were found to react strongly with Sh. flexneri, S. typhimurium and S. enteritidis. The Natal Blood Transfusion Service has found that in Natal, blood units containing high concentrations of specific antibodies occur with a frequency of 3,6% among all White donors and 10,35% among all African donors. They found that African females, in turn, had almost twice the frequency of high titre serum as African males. In this study, Indian female hospital patients did not have a statistically higher frequency of high-titre serum than Indian male patients. Blood units donated to the Natal Blood Transfusion Service are now routinely screened by ELISA for antiendotoxin antibodies and those units with high concentrations (> 40 ug/ml) of antibody were pooled and fractionated to obtain a gamma globulin, Lot LG-l. The binding capacity of the LG-1 antibodies towards 12 endotoxins was examined. Binding was found to be highest with endotoxin from Sh. flexneri, S. abortus equi and S. typhimurium and intermediate with S. enteritidis and E.coli 026:B6. Binding with the other endotoxins tested was relatively low. Differential absorption experiments showed that LG-1 was made up of a mixture of cross-reacting as well as specific antibodies For example, the antibodies binding Sh. flexneri endotoxin were mainly specific. Those binding E. coli 026:B6 endotoxin were specific and cross-reacting in almost equal proportions. Antibodies to the endotoxins from the salmonella strains tested were mainly cross-reacting. The specificities of the LG-1 antibodies towards endotoxins from the various Gram-negative bacteria did not in most cases reflect the incidence of these organisms in blood cultures taken from hospital patients. V The activity of LG-1 antibodies was compared to that of normal human immunoglobulin preparations obtained from the National Blood Fractionation Centre, Pinetown and to an anti-pseudomonas immunoglobulin prepared by Wellcome Laboratories, England. The binding capacity of the antibodies in the standard globulin preparations towards most of the endotoxins tested was less than 15% of that of the LG-1 antibodies. The anti-pseudomonas immunoglobulin was shown to bind poorly to most of the endotoxins tested in comparison with binding by LG-1 antibodies.
The impact of social identity and selected other variables on the career maturity of a small sample of coloured matriculants at high schools in Pietermaritzburg.
(1995) Freeman, Suzannah Rosemary.; Basson, Clive James.
Abstract available in PDF.
Determination of exposure of humans to selected mycotoxins with particular reference to aflatoxins.
(1995) Early, Deborah Angeline.; Dutton, Michael Francis.; Chuturgoon, Anil Amichund.
Mycotoxins are poisonous secondary metabolites commonly produced by fungi and are involved in human disease conditions known as mycotoxicoses. There is evidence to show that food eaten by the rural Black population of Southern Africa is contaminated with mycotoxins. A tenuous relationship exists between the occurrence of mycotoxins in foods and certain disease conditions in humans. In order to verify this relationship, efforts have, in the past, been made to detect mycotoxins and their metabolites in physiological fluids and tissues. The difficulty with this approach is that mycotoxins in the body have short half lives, being rapidly excreted or metabolised to other forms. More recently it has been shown that aflatoxin B1, as its activated epoxide, can conjugate with macromolecules such as nucleic acids and proteins. These survive for much longer than the free toxins and by suitable methods can be isolated and measured. This allows for a much better estimate of exposure of the individual to aflatoxin. This study reviews and evaluates screening methods for the detection and analysis of mycotoxin contamination in rural foodstuffs such as maize and groundnuts. Methods for the production of aflatoxin-lysine and protein adducts are motivated and developed then used in the identification of naturally occurring adducts in humans. Isolation and quantitative analysis techniques are proposed to routinely screen patients for evidence of aflatoxin exposure.
A cytotoxic evaluation of aflatoxin B1, zearalenone and their epoxide derivatives using human cell lines.
(1996) Pillay, Dharmarai.; Chuturgoon, Anil Amichund.; Dutton, Michael Francis.
Since the discovery of mycotoxins in food, the thrust of biochemical and toxicological research has been carried out on animals which has proven to be uncoordinated and not easily extrapolated to humans. Over the last decade, there have been increasing pressures to review and reduce the use of animals in experimental toxicological studies. Consequently in this study aflatoxin B1 (AFB1), zearalenone (Zea) and their epoxide derivatives have been evaluated using in vitro assays. The HepG2, A549 and Hela cell lines were used for assessing the cytotoxicity, effects on cellular metabolism and sites of action of AFB1, Zea and their derivatives. The cytotoxicity of these mycotoxins was evaluated using the methylthiazol tetrazolium (MTT) reduction assay. Cells, treated with mycotoxins were prepared for transmission electron mlcroscopy (TEM), immunocytochemistry (ICC), scanning electron microscopy (SEM), confocal and light microscopy. From the cytotoxicity assay it was found that the epoxide derivatives were more toxic than the parent toxin when exposed to HepG2 cells with no significant differences in toxicity levels in A549 and Hela treated cells. Both epoxide derivatives displayed a regression of hepatoma cell proliferation at high doses (25ug/ml) while lower concentrations (<12.5ug/ml) enhanced cell growth. Microscopy analyses showed distinct cellular alterations. When exposed to AFB1 (12.5ug/ml) hepatoma cells showed prominent ultrastructural alterations such as areas of cytoplasmic lysis and increased numbers of secondary lysosomes while cells exposed to Zea (l2.5ug/ml) displayed numerous ovoid mitochondria and proliferation of rough endoplasmic reticulum which is indicative of enhanced protein synthesis. The presence of label in toxin treated cells is suggestive of the effects of these mycotoxins. Such cellular changes may lead to altered metabolism and cell function.
Evaluation of the anthropometric parameters and fitness levels of prepubertal Indian soccer players.
(1997) Jagot, Mahmood Abdull Rahim.
Due to the lack of morphological data on prepubertal Indian male soccer players in South Africa, this study was undertaken on ninety male prepubertal subjects. The subjects were divided into three groups of thirty subjects each: Experienced "E" (those playing organized soccer for more than two years), beginners "8" (those playing organized soccer for less than two years) and sedentary "S" (those not participating in organized soccer). All subjects were measured according to Heath - Carter anthropometric somatotype methods. Fitness tests comprising power and strength tests (vertical jump height and standing broad jump) and muscle endurance tests (push - ups and sit - ups) were also done. The three groups were first compared to each other and then to available international data. There were no statistical differences among the three groups for: height, weight, age, triceps, subscapular, suprailiac, calf and total skinfolds, humerus and biceps girth, ectomorphy, mesomorphy and endomorphy, suggesting a general homogenicity between groups. For fitness tests the "E" group performed significantly better than the others for standing broad jump and sit - ups (p = 0.005 and p = 0.036 respectively). For push - ups the "8" and "E" were significantly better than the "S" group, (p = 0.013, for "8" versus "S" group), indicating that in soccer muscle strength and explosive strength are important. The lack of difference between the groups for anthropometric criteria in this study may be explained by the experienced players' inadequate training. Other factors may include the lack of parental involvement, inadequate knowledge on fitness aspects and poor training methods. Furthermore, the sedentary group may be participating in unorganized activities which renders them at a level similar to the experienced group. Data on non - Indian South African junior players is required to help us understand the lack of significant Indian talent in the National team. Other factors such as diet, cultural differences, training methods, level of coaching, environmental factors and sport facilities need investigation and be addressed if we want to see an improvement in the South African Indian soccer players.
Heart rate response and ECG monitoring in veteran squash players.
(1997) Sibbald, Helen.
The incidence of sudden death during or after squash play has become a source of concern. In order to screen for coronary artery disease, exercise stress testing has been advocated, by the American College of Sports Medicine (1986), for those at or above the age of 45 already exercising or before embarking on exercise. Eighteen veteran squash players (mean age 49 ± 3 yr) took part in the study. Heart rate response was monitored throughout a squash match and for an hour after play. ECG changes were monitored for one hour after squash play. Mean heart rate, throughout playing time was 148 ± 16 beats per minute (range 118 - 168 bpm), representing 86.7% of Predicted Maximum Heart Rate (PMHR). Mean maximal heart rate was 169 ± 14 bpm (range 141 - 186 bpm), representing 98.8% of PMHR. Thus squash represents a very high intensity activity for these players. On subsequent ECG monitoring, no abnormalities were detected. The results of this study confirm that squash is an extremely high intensity sport and that even veteran players play at a level close to their maximal. This level of play did not provoke subsequent cardiac arrhythmias in this small group of players, contrary to an earlier study that reported arrhythmias in one third of a group of younger players in the post match period.
The effect of infra-red laser therapy on fibroblast activity in cell culture.
(1997) Mars, Susan.; Chuturgoon, Anil Amichund.
Abstract available in PDF.
The use of laser doppler fluxmetry in the pre-operative assessment of amputation wound healing in the dysvascular patient.
(1997) McKune, Andrew James.; Mars, Maurice.; Robbs, John Vivian.
Abstract available in PDF.
Mycotoxins in food with particular reference to fumonisin B1 : their health impact on a Kranskop rural community, KwaZulu Natal.
(1998) Chelule, Paul Kiprono.; Dutton, Michael Francis.; Gqaleni, Nceba.
The use of the multi-mycotoxin screen based on dialysis to analyze foods and feeds for mycotoxins, is well documented. This study investigated the possibility of incorporating FB I into the screen. Maize meal (25g) was spiked with AFB I , CPA, FB1, ST and ZEA and extraction was done using acetonitrile/4% potassium chloride (90:10 v/v). The recoveriesof the mycotoxins were 77.4, 61.5, 97.4, 79.8 and 98% respectively on analysis by HPLC. Fumonisin B1 could not be completely incorporated into the screen due to its reaction with sodium hydrogen carbonate, which is a component in the method. Thus, FB I was determined in a separate portion of the extract. The high cost of FBI standards which are often of inferior purity necessitated that FB I standards be locally produced in the laboratory using Fusarium moniliforme MRC 826, a good producer of FB 1 . In this study, production of FB I was carried out using a stirred jar fermenter and patty cultures. The yields were 160mg/1 and 6mg/g of FB I for the two methods respectively. Methyl esterification of tricaballylic acid moieties of FB I was done for effective clean-up. This was achieved by derivatizing FBI, with diazomethane. It was found that other functional groups besides the tricaballylic acid moieties of FB I were undesirably methylated as well, which made cleanup by this method difficult as shown by electrospray mass spectrometric analysis. Attempts to de-methylate FBI methyl esters with esterase was not successful. Analysis of human faecal samples was carried out with the view of developing a short term marker for assessing human exposure to FB I . Faeces from rural (20) and urban (23) volunteers were analyzed by high performance liquid chromatography. The results showed that 35% of the rural samples and 9% of the urban volunteers had detectable amounts of FB I ranging from 0.600 to 19.56 mg/kg. There was a significant difference (p = 0.04)between the two population groups. A study was carried out to assess the occurrence of FBI in a rural area of Tugela valley in Kranskop magisterial district of KwaZulu Natal. A questionnaire was administered to gather information on the family health and nutrition. Raw (stored) and processed foods and faeces, were collected for analysis of FB1. A similar control study was carried out in the urban area of Durban Metro. Homes were mapped out using the GIS for easy follow up. Oesphageal cancer (OC) incidence from the local hospital and weather data for the study area were collected from South African Weather Bureau, Johannesburg. The questionnaire results showed that the common diseases were mainly of respiratory origin (24% and 26%) from both rural and urban groups respectively. Food analysis (by HPLC) showed that the number of maize samples with FB I were higher in the rural area (31.9%) in comparison to the urban samples (6.1%). The level ranged from 0.092-22.225 mg/kg in food and 0.513-39 mg/kg in faeces. The mean concentration of FB i in the faeces and maize samples showed a similar significant difference of 0.014 between the two groups. However, these concentrations were much lower than those of high OC area in Transkei (117 mg/kg). There was no detection of FBI in fermented food products.
The occurrence and detection of aflatoxin-macromolecular conjugates in humans.
(1998) Myeni, Sibongiseni Selby.; Chuturgoon, Anil Amichund.; Dutton, Michael Francis.
Aflatoxin Bi (AFBi), a highly toxic fungal metabolite (mycotoxin) of certain strains of Aspergillus, has long been known to be carcinogenic in animal species. Accumulation of epidemiological evidence led to its classification, in 1993, by the International Agency for Research on Cancer as a Group I human carcinogen. Aflatoxin Bi contaminates the food supply in most tropical and sub-tropical countries, where it is associated with increased incidence of hepatocellular carcinoma (HCC). In these countries, AFBi is also linked to kwashiorkor, jaundice, and Rey's syndrome. The biological action of AFBi is through its oxidation to AFBi-8,9 epoxide (AFBiO). This epoxide binds to macromolecules like DNA, RNA and proteins as well as amino acids to form AFBi-macromolecular adducts. Quantitation of these adducts is thought to be the most promising approach in the development of methods to measure levels of exposure to aflatoxins. Aflatoxin Bi was produced, isolated and purified using preparative thin layer chromatography (TLC). The toxin was oxidised to AFBiO using dimethyldioxirane and the UV spectra of both the AFBi and AFBiO were determined. Reaction of selected Na-acetyl amino acids (AA) with AFBiO was studied and UV spectrophotometry, TLC, high performance liquid chromatography (FfPLC) and high performance capillary electrophoresis (CE) were used to characterise the reaction products. The epoxide was also reacted with albumin and DNA. Aflatoxin Bi-albumin reaction mixture was hydrolysed and characterised by TLC. Spectrum measurement of the oxidative product of AFBi gave peaks at 266 and 367nm. Qualitative TLC and the epoxide spray reagents confirmed that epoxidation was successful. The in vivo reaction of selected Na-acetyl AA with the epoxide gave peaks between 300 and 400 nm. Naacetyl-arginine, Na-acetyl-lysine and Na-acetyl-histidine showed reaction with AFBiO with maximum wavelengths at 392, 397 and 391 nm respectively. These results strongly suggest that AFBiO is able to covalently bind to lysine, histidine and arginine in albumin. A total of twenty nine blood samples were analysed by HPLC for the presence of AFBilysyl adduct. Of the twenty nine samples, ten were from HCC patients, ten from control patients and nine from kwashiorkor patients. The results show that AFBi-lysine does occur in patients at King Edward VIII Hospital (KEH) and the highest level was detected in HCC patients followed by kwashiorkor patients.
The immunocytochemical and electrophoretic localisation of aflatoxin B1-binding proteins in isolated liver mitochondria.
(1998) Raman, Gareth.; Chuturgoon, Anil Amichund.; Dutton, Michael Francis.
Mitochondria perform functions which are central to the life of most eukaryotic cells. These organelles can be considered the ultimate energy power house of a living cell. The role of mitochondria in cancer phenotype remains a fertile area of research. Several carcinogens are known to enter the mitochondria, resulting in impaired functioning and altered structure. Aflatoxin BI (AFB1) a primary type I mycotoxin elaborated by Aspergillus flavus and Aspergillus parasiticus, is carcinogenic for a wide species range. The epoxide is capable of binding to nucleic acids and proteins, resulting in induced mutations, cellular toxicity, and eventually carcinogenesis. Approximately 250 000 deaths occur annually in both China and Africa due to patients presenting with Hepatocellular Carcinoma (HCC). The causative agents being AFB1-ingestion via contaminated foods and feeds, and the Hepatitis B Virus infection. The toxin has a multifaceted mode of attack, capable of being activated to a highly reactive and carcinogenic derivative, the AFB1-8,9-epoxide, via the cytochrome P450 enzyme system of the microsomes, endoplasmic reticulum and also the mitochondria. The epoxide is capable of binding to nucleic acids and proteins, resulting in the formation of covalent adducts. The repeated occurrence of gold labelled toxin within mitochondria from hepatomas of patients presenting with HCC suggested that these organelles were direct sites of toxin binding. Despite observations that mitochondria appear as direct and perhaps preferential targets for attack by AFB1, the actual in vivo immunolocalisation and characterisation of bound AFB1 within liver mitochondria has not been reported previously. In addition the role of AFB1-protein binding within mitochondria was investigated to determine the mode of action of the toxin, within the mitochondrial system. Liver sections from rats treated with a single lethal dose of AFB1, showed distinct ultrastructural abnormalities viz. large nuclei, increased heterochromatin, and swollen mitochondria. Immunocytochemistry revealed for the first time, the selective localisation of conjugated gold labelled toxin within the mitochondria. Toxin was found in the intracristal and peripheral spaces and frequently within the mitochondrial matrix. The mitochondria isolated from treated rats revealed significant alterations and damage to the mitochondrial membranes. The cristae were also markedly swollen with the associated clearing of the mitochondrial matrix. Western blot immunoassays revealed the presence of five AFB1-bound proteins (150kDa, 50kDa, 25kDa, 18kDa, 14kDa) in the inner mitochondrial fraction of isolated mitochondria. High pressure liquid chromatography also revealed that a significant proportion (84%) of an initial dose of toxin, was absorbed by mitochondrial protein. This study is the first to show the presence of specific mitochondrial proteins involved in toxin binding. In addition, the presence of toxin within the mitochondria and the specific binding to inner mitochondrial proteins suggest that the toxin specifically targets the electron transport chain and hence effects ATP production. This study conclusively indicates that mitochondria are direct targets for attack by AFB1 during experimental carcinogenesis. Mitochondria therefore play an important role in AFB1-mediated carcinogenesis.

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