Browsing by Author "Smith, Michael Trevor."

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Alkaloids of Catha Spp.
(2001) Field, Courtney Robin.; Smith, Michael Trevor.
The levels of the psychoactive alkaloids S-(-)-cathinone and its primary metabolite cathine, consisting of the diastereomers (+) -norpseudoephedrine and (-)- norephedrine were determined in Catha edulis (Vahl) Forssk. ex Endl., Catha transvaalensis Codd and Catha abbottii Van Wyk & Prins. Alkaloid levels were investigated in C. edulis plants collected from three different localities in South Africa, and one from a Nairobi khat market. The efficiency of three different methods for the extraction and isolation of cathinone and cathine were investigated, viz. an aqueous acid extraction, an organic solvent extraction and an aqueous acid extraction using the commercially available Extrelutᴿ procedure. The aqueous acid extraction resulted in the rapid loss of cathinone and yielded variable alkaloid levels in replicate studies. This was also observed when this method was coupled with the Extrelutᴿ procedure. In contrast, the organic solvent extraction did not result in a loss of cathinone and provided consistent results over a number of replicates; it also proved to be a simple and rapid method for extracting and isolating cathinone and cathine. A trifluoroacetic acid (TFA) derivatization procedure which has been suggested to produce characteristic diagnostic fragments for gas chromatography / mass spectrometry (GC/MS) identification, was investigated, but failed to produce consistent TFA derivatives of cathinone and cathine. However, underivatized cathinone and cathine were easily identified by GCMS due to their unambiguous mass spectra. All subsequent studies were undertaken using the organic solvent extraction and isolation method, coupled with GC analysis and GC/MS identification of underivatized cathinone and cathine. Leaves of C. edulis were found to contain cathinone and cathine at levels 100 times higher than those of C. transvaalensis. The alkaloids were undetectable in C. abbottii. Plants grown from cuttings of C. edulis collected from the Durban Botanical Gardens were found to contain cathinone and cathine at levels of 0.410 mg per gram fresh weight and 0.157 mg per gram fresh weight in leaves, respectively, while these levels in plants derived from different localities decreased in the order: Eastern Cape (0.319 mg/g f.w cathinone and 0.029 mg/g f.w cathine), Mpumalanga (0.139 mg/g f.w. cathinone and 0.171 mg/g f.w. cathine) and Nairobi (0.032 mg/g f.w. cathinone and 0.025 mg/g f. w. cathine). In an investigation of the cathinone levels in the different plant parts it was found that the highest levels were found in leaves of the shoot tip (0.243 mg/g f.w.) but decreased with the age of the leaf and developmental stage of the plant in the order: juvenile leaves (0.124 mg/g f.w.), mature leaves (0.035 mg/g f.w.), young stem (0.033 mg/g f.w.) and mature stem (0.004 mg/g f.w.). Concomitantly, cathine levels increased with the age of the leaf: leaves of the shoot tip (0.006 mg/g f.w.), juvenile leaves (0.011 mg/g f.w.), mature leaves (0.019 mg/g f.w.). The cathine level in the young stem material was found to be the highest in the entire plant (0.270 mg/g f.w.) but decreased markedly in the mature stem (0.052 mg/g f.w.). Both cathinone and cathine levels in the mature root were greater than levels in the mature stern, being 0.012 mg cathinone per gram fresh weight, and 0.063 mg cathine per gram fresh weight. Neither cathinone nor cathine were detectable in young root material.
Biochemical and ultrastructural changes associated with chilling injury in soybean seeds during imbibition.
(1996) Roskruge, Carol Lynette.; Smith, Michael Trevor.
Biochemical and ultrastructural changes associated with chilling injury (CI) in soybean seeds imbibed at 5°C and 25°C were investigated. Soybean seed germination appeared to be affected by chilling temperatures and initial seed moisture content. Seeds with higher moisture contents exhibited 85% germination, while low moisture content seeds had a 32% germination. Leakage rates were greater in chilled seeds, indicating that membrane integrity in the tissues was impaired at chilling. The low rates of potassium ion leakage between 6 and 24 hours of imbibition compared to the high peroxide levels observed during this period led to the suggestion that lipid peroxidation was a better marker of CI than leakage. Transient changes in lipid hydroperoxide levels were observed in chilled and non-chilled seeds and axes. However, in axes, the increase in lipid hydroperoxides after 12 hours of imbibition at chilling temperatures was associated with an 18% decline in linoleic acid levels of total lipid fraction. Similarly, a 10% decline was observed in the polar lipid fraction. These results suggest that the capacity of seeds to control lipid peroxidation may be an important component in CI and that a consequence of peroxidation is likely to be a loss of fatty acid unsaturation. Sugar levels were not affected by chilling and non-chilling temperatures and no relationship could be established with CI. Antioxidant defense enzymes (catalase and superoxide dismutase) were expressed at chilling and non-chilling temperatures and increases were observed after 24 hours of imbibition which showed an apparent correlation with increases to lipid hydroperoxide levels. Enzyme levels decreased after 48 hours of imbibition at a time which coincided with the decline observed in the peroxide levels. Overall, no marked differences were observed in chilled and non-chilled cells at the ultrastructural level, except that vacuolar reserve mobilization was markedly impeded.
Detecting parasites loads in urine diversion toilets.
(2009) Hawksworth, David James.; Smith, Michael Trevor.; Rodda, Nicola Heike.
In an attempt to supply sanitation to the growing communities in rural and peri-urban areas around Durban, the eThekweni Municipality has installed urine diversion (UD) toilets which have been modified to suit local conditions . These toilets are based on the ecological sanitation (EcoSan) system. The future aims are to reuse waste as a composting medium and minimize the use of water but the presence of microorganisms in the faecal waste poses a potential health risk to people in contact with it. Currently the Municipality has not deemed the waste safe for re-use but has suggested that after a one year standing period it should be free of all potential pathogens including Ascaris lumbricoiodes (human roundworm) ova. This study reports on the development of the AMBIC protocol for the recovery of Ascaris ova from the standing vaults of UD toilets. The protocol has been shown to consistently recover over 70% of Ascaris ova and has the added advantage of recovering the ova of other helminth species (Trichuris trichiura and Taenia sp.) present in a UD standing vault sample. Recoveries of Ascaris ova and ova of other parasite species, namely Trichuris and Taenia sp., are reported from waste which has been standing for one year. This is cause for concern as it shows one year is not a sufficient standing period to render the waste free of all microorganisms. Sampling from 124 UD toilet vaults that were in use, showed a high prevalence of both helminth (Ascaris lumbricoiodes, Trichuris trichiura and Taenia sp.) and protozoan (Giardia and Cryptosporidium) parasites.
The financial and economic feasibility of biodigester use and biogas production for rural households.
(2011) Smith, Michael Trevor.; Blignaut, James N.; Goebel, Jessica Schroenn.
In South Africa, sustainable development is set in the context of two separate economies. The second of these economies consists of the rural population and is characterised by poverty and stagnant development. Sustainable development is an increasingly topical concept which highlights the need for development to proceed in a manner that does not deplete natural resources. In addition to narrowing the gaps between the various classes (layers) in an economy, the key ‘ingredients’ of sustainable economic development include “natural resource management, food, water, and energy access, provision and security” (Blignaut, 2009: cited in Blignaut and van der Elst, 2009: 14). A biodigester is a potential solution to some of the difficulties faced by remote rural populations. Biodigester systems are submerged tanks capable of producing a nutrient rich fertiliser and combustible gas when consistently fed with organic matter and water. A biodigester may be one simple answer to the key ingredient needs of sustainable development – reducing the depletion of natural resources, providing clean burning energy for cooking and fertiliser for growing food. The potential is clear for biodigesters to aid in the process of sustainable development. The question to be analysed is whether this technology would be financially and economically feasible for installation and use in rural households. This thesis focuses on a typically remote and rural community in KwaZulu-Natal, South Africa, in order to assess the potential feasibility of a biodigester system. The appraisal takes the form of a Cost Benefit Analysis (CBA) and aims to establish whether or not this technology is financially feasible for individual rural households and/or economically beneficial to society.
The health-related microbial quality of drinking water from ground tanks, standpipes and community tankers at source and point-of-use in eThekwini Municipality : implications of storage containers, household demographics, socio-economic issues, hygiene and sanitation practices on drinking water quality and health.
(2009) Singh, Urisha.; Smith, Michael Trevor.; Rodda, Nicola Heike.
The aim of this study was to investigate the microbiological quality of drinking water at the source (taps at eThekwini laboratories, standpipes and mobile community tankers) and corresponding point-of-use (storage containers and ground tanks) supplied to peri-urban areas in Durban by eThekwini Municipality. It also aimed to identify factors associated with deterioration in water quality such as storage of water, household demographics, hygiene and sanitation practices. In order to determine the microbial quality of drinking water, the pour plate method (for enumeration of heterotrophic organisms) and the membrane filtration technique (for total coliforms and E. coli enumeration) were used. Conductivity, turbidity, pH and total and residual chlorine levels of drinking water were measured. Microbial and physico-chemical data was collated and statistically analysed with epidemiological data from an associated study to determine the link between microbial quality of drinking water, household demographics, health outcomes, socio-economic status, hygiene and sanitation practices. Findings showed that all point-of-use water was unsafe for human consumption as a result of either poor source water quality, in the case of standpipes, and microbial contamination at the point-of-use, in the case of ground tanks and community tankers. The latter could be attributed to unsanitary environments, poor hygiene practices or poor wateruse behaviour. Households which included children aged 0-5 years and in which open-top containers were used for water storage had the highest rates of diarrhoea and vomiting. Water from ground tanks had the best microbial quality but people in households using this water presented with the highest rate of diarrhoea. Therefore provision of microbially safe drinking water will not reduce the rate of health outcomes if addressed in isolation. In order to reduce water-associated illness, provision of safe and adequate amounts of water, hygiene and sanitation education and education on water-use behaviour should be provided as a package. The provision of improved water delivery systems does not ensure that drinking water is safe for human consumption. Measures, such as point-of-use water treatment should be considered to ensure that drinking water provided at the source and point-of-use is microbially safe for human consumption.
In vitro polyploidization of selected indigenous plant species.
(2005) Reddy, Viloshanie.; Smith, Michael Trevor.
Many plant species indigenous to South Africa have ornamental, medicinal and horticultural value. Polyploidization is one technique that has been used to artificially produce superior genotypes, particularly in horticultural species. In the current investigation two antimitotic substances, colchicine at concentrations of 0.1% and 0.01% and oryzalin at concentrations of 0.01% and 0.001%, were used in an attempt to polyploidize microshoots of Dorotheanthus bellidifonnis (Burm.f.) N.E.Br and Mondia whiteii (Hook.f.) in vitro. Microshoots of D. bellidifonnis and M. whiteii obtained from nodal cuttings of in vitro germinated seedlings were maintained for 48 hours in liquid medium containing the antimitotic substances and thereafter cultured on sucrose-supplemented MS medium. The treated microshoots were evaluated for elongation, necrosis, contamination and phenolic exudation. Best results were observed in M. whiteii microshoots treated with antimitotic substances and transferred on to solid sucrose-supplemented MS medium containing 0.2% activated charcoal for 4 weeks. Leaves from the surviving treated plants were excised and used for flow cytometric analyses to evaluate changes in chromosome number. Shoots of M. whiteii treated with 0.01 % colchicine showed no changes in chromosome number, while the higher concentration used produced polyploids and mixaploids. However, oryzalin at 0.01 % concentrations produced a comparatively higher number of microshoots that were polyploids and mixaploids. Shoots of M. whiteii that have altered chromosome number have been transferred onto multiplication medium, for future evaluation of changes in phenotypic characteristics. The germination response of seeds of D. bellidifonnis was evaluated in the presence of oryzalin (0.01% and 0.001%) and colchicine (0.1% and 0.01%). Poor germination was observed in seeds germinated in the presence of 0.01 % oryzalin. Upon transfer of the germinated seedlings treated with antimitotic substances onto sucrose-supplemented MS medium, subsequent growth and development was restricted. Shoot and root development was different for the seedlings germinated in the presence of the two antimitotic substances. Shoot elongation and root development was vigorous in seedlings geminated in the presence of 0.01 colchicine and stunted development was observed in seedlings germinated in 0.1 % colchicine. On the other hand restricted rootind was observed in seedlings germinated in the presence of oryzalin, and the shoots lacked pigmentation. Meristematic cells excised from the shoot tips of the treated seedlings showed several ultrastructural changes including abnormal mitochondrial development, endomembrane formation and vacuolation. It was concluded that oryzalin and colchicine influence ultrastructure in plant cells differently. Practical constraints associated with plant tissue culture also influence the rate of in vitro polyploidization. Since different plant species require different conditions for optimal growth, it was also noted that no unique polyploidization treatment can be used for a wide range of plant species, individual species require different growth conditions.
Lipid peroxidation and ageing in seeds of cabbage and soya bean.
(1986) Hailstones, Milson Donald.; Smith, Michael Trevor.; Campbell, Glen Kerry.
It has been suggested that lipid peroxidation is involved in the loss of seed vigour, although many attempts to examine the relationships between lipid peroxidation and seed vigour have proved equivocal. Studies were undertaken on seed lots of cabbage and soya bean to find evidence for peroxidation by the analysis of i) total and polar fatty acid levels; ii) lipid hydroperoxides; iii) volatile products produced on heating dry seeds; and iv) volatile products produced on imbibition. The loss of polyunsaturated fatty acids (PUFAs) detected in the dry seeds was clearly related to germinability in both soya bean and cabbage seeds. Furthermore, an increase in hydroperoxides was observed in both seed types. Although the relationship of the level of hydroperoxides to germinability was less clear than for the decline in the level of PUFAs, these results suggested that the loss of PUFAs was possibly due to evidence the peroxidation of the seed obtained from the heating lipid, indirect of the seeds suggesting that hydroperoxide breakdown may be necessary in order that the changes in PUFAs become apparent. In contrast to the poor relationship observed between germinability and hydroperoxide level, a marked relationship between hydroperoxide level and seed moisture content was observed in the cabbage seeds. This may be significant with regard to the observed relationship between storability and seed moisture content, although no such relationship was seen in the soya beans. Certain volatile compounds derived from dry heated seeds were related to seed vigour in both seed types and evidence suggests that the lipid hydroperoxides were the source of these compounds. Although the total volatiles counts evolved from imbibing cabbage seeds showed no quantitative relationship to seed vigour, one peak was noted which was clearly associated with the vigour of these seeds. The variability in the volatiles evolved from soya beans on imbibition, however, precluded the detection of any possible relationship between these and seed vigour. In both seed types, results suggest that the volatiles derived on imbibition were of a different source to those derived on heating. A marked increase in the level of hydroperoxides was observed in whole cabbage seeds and soya bean axes of low vigour over the first hour of imbibition. This may suggest that an exacerbation of damage on imbibition was associated with low vigour seeds. In contrast to this, in the seeds of high vigour, the increase in hydroperoxide levels was markedly less or rapidly reduced, suggesting the possible activity of repair mechanisms. Ferrous ions were shown to invigorate both seed types, particularly cabbage seeds. It is suggested that the invigorating effect of these compounds was due to the facilitation of repair, including hydroperoxide breakdown and the quenching of any free radicals.
Lipid peroxidation and ageing in seeds of Glycine Max.
(1990) Hailstones, Milson Donald.; Smith, Michael Trevor.; Campbell, Glen Kerry.
Six different lots of soya beans (Glycine max (L.) Merr.) were examined. Seed hydroperoxide levels were highly correlated with viability, but not with moisture contents. It was proposed that moisture contents may exert a similar antioxidant effect at intermediate levels as has been observed in dry foods. Seeds treated with ferrous sulphate were significantly (S% level) invigorated. Furthermore, this treatment was observed to give rise to a reduction in the peroxide value of soya bean axes over the first hour of imbibition, an increase in 2,3,S-triphenyl-tetrazolium chloride reduction and protein synthesis, and a decline in electrolyte leakage. It was proposed that this was due to the antioxidant activity of the ferrous iron, leading to an attenuation of free-radical induced autoxidation. Ferrous sulphate treated seeds produced more aldehydes than untreated seeds. This result suggested that aldehydes may not be responsible for declining seed vigour. Hexanal, pentanal and butanal production from heated dry seeds was significantly correlated with seed germination, CVG and hydroperoxide levels. The thermal breakdown of the hydroperoxides was postulated to be the source of these compounds. A GC technique was developed using model systems of oxidized methyl oleate, linoleate, linolenate and soya bean bulk oil. The analysis of seed lipid oxidation products revealed marked differences in the proportions of the products compared to bulk and monolayer oxidation. The selective production of the 13-hydroperoxide implicated enzymatic or metalloprotein involvement. The implications of the results of this study with regard to the present theories of seed ageing were discussed.
Lipid peroxidation and the antioxidant systems in soybean seed maturation and germination.
(2003) Tyiso, Sakiwo.; Smith, Michael Trevor.
The biochemical changes taking place during soybean seed development and gennination, and some aspects of desiccation tolerance were assessed with reference to lipid peroxidation and antioxidant systems. During nonnal seed development, fresh weight and dry weight increased between 20 and 50 days after flowering (DAF), concomitant with the accumulation of triacylglycerols and sugar reserves, after which dry weight remained almost unchanged, and fresh weight decreased. Seed moisture content decreased rapidly during the last stages of development. High levels of lipid peroxidation were evident between 20 and 45 DAF, and decreased thereafter. An examination of antioxidant systems revealed that whereas total glutathione levels accumulated continuously throughout the 80 days of seed development, both dehydroascorbic acid (DHA) reductase and ascorbate free radical (AFR) reductase increased concurrently with the increase in total ascorbate content, and the overall levels did not decrease markedly during maturation drying. Ascorbate peroxidase (ASC POD) activity was high during the period ofgreatest ascorbate accumulation. Both catalase (CAn and superoxide dismutase (SOD) activities increased progressively during early seed development (20-40 DAF), but showed variable patterns of change during maturational drying, in marked contrast to ASC POD which declined from 40 DAF to undetectable levels at 70 DAF. An assessment of the relationship between the antioxidant systems and lipid peroxidation was made during imbibition and gennination, as it has been suggested that controlling free radicals was a critical event in early imbibition. Unexpectedly, lipid peroxidation increased progressively in both seeds and isolated axes, and were eight-fold higher at 48 hours of imbibition compared to dry tissues. A progressive, and co-ordinated, increase in CAT, total glutathione, total ascorbate pool, guaiacol POD, ASC POD, and SOD appeared to parallel the rise in lipid peroxidation in both whole seeds and axes. Variable responses were evident between seeds and axes for the enzymes AFR reductase and DHA reductase In order to gain a further insight into the dynamics of desiccation-tolerance and desiccationsensitivity, imbibing seeds were subjected to an unscheduled dehydration treatment, and then rehydrated for up to 24 hours. During these hydration-dehydration-rehydration (H-D-R) treatments, changes in lipid peroxidation and antioxidant systems were measured. Concurrent with the loss of viability in the axes of seeds dehydrated after 24 and 36 hours of imbibition, there were increases in both lipid peroxidation and solute leakage. Unscheduled drying was seen to be a critical stage, as intolerant axes showed four- to eightfold increases in lipid peroxidation, which were only partially reduced on subsequent rehydration. Tolerant axes, on the other hand, were able to maintain low, basal levels of lipid hydroperoxides on drying. The relationship between these observations and the antioxidant systems showed that the antioxidant enzymes CAT, ASC POD, AFR reductase, DHA reductase, guaiacol POD and SOD declined markedly during the unscheduled drying, whereas GSH and ASC declined only slightly. On rehydration, most of the enzymes, total glutathione, and total ascorbate pool increased, the only exception being the loss of ASC POD activity. ORA reductase, which was seen to decrease as a part of nonnal gennination, declined progressively also in H-D-R treatments. These results suggested that loss of viability was not attributable to a decline of the antioxidant systems but rather to the combined deleterious effects of increased lipid peroxidation, and a generalized and moderately compromised antioxidant system. These studies have indicated that the occurrence of lipid peroxidation can be seen as a nonnal part of seed development and gennination. The H-D-R studies, on the other hand, supported the concept that the balance between peroxidation reactions and the protective systems was critical to the development of desiccation tolerance.
Membrane changes and lipid peroxidation during ageing in seeds of Lactuca sativa L.
(1986) Smith, Michael Trevor.; Campbell, Glen Kerry.
Abstract available in PDF file.
A phytochemical investigation of two South African plants with the screening of extractives for biological activity.
(2006) Gallagher, Andrew Bruce.; Koorbanally, Neil Anthony.; Smith, Michael Trevor.
Two South African medicinal plants, Strophanthus speciosus and Eucomis montana, were investigated phytochemically. From Strophanthus speciosus a cardenolide, neritaloside, was isolated, whilst Eucomis montana yielded three homoisoflavanones, 3,9- dihydroeucomin, 4'-demethyl-3,9-dihydroeucomin, and 4'-demethyl-5-0-methyl-3,9- dihydroeucomin. The structures were elucidated on the basis of spectroscopic data. The homoisoflavanones were screened for anti-inflammatory activity using a chemiluminescent luminol assay, modified for microplate usage. All of the homoisoflavanones exhibited good inhibition of chemiluminescence, with IC50 values for 3,9-dihydroeucomin, 4'-demethyl-3,9-dihydroeucomin, and 4'-demethyl-5-0-methyl-3,9- dihydroeucomin being 14mg/mL, 7mg/mL, and 13mg/mL respectively. The IC50 value of 4'-demethyl-3,9-dihydroeucomin compared favourably with the NSAID control (meloxicam), which had an IC50 of 6mg/mL. Neritaloside was not screened for biological activity as the yield of 14.4mg was insufficient for the muscle-relaxant screen for which it was intended. An assay for antioxidant/free radical scavenging activity was also performed. All the compounds had excellent antioxidant/free radical scavenging activity, with percentage inhibition of the reaction being 92%, 96%, and 94% for 3,9-dihydroeucomin, 4'-demethyl- 3,9-dihydroeucomin, and 4'-demethyl-5-0-methyl-3,9-dihydroeucomin respectively at a concentration of 10mg/mL. However, the control compounds, diclofenac and meloxicam, also exhibited strong activity, with the result that the precise mode of anti-inflammatory activity could not be unequivocally determined. The results from the biological screenings thus provided a rational scientific basis for the indigenous ethnomedicinal use of Eucomis species in the treatment of rheumatism, inflammation and pain.
Post-phloem transport and metabolism of sucrose in avocado.
(2001) Cripps, Ryan F.; Smith, Michael Trevor.; Cowan, Ashton Keith.
In South Africa, and in several other sub/tropical countries, the avocado represents a commercially important crop. Very little is currently understood about the metabolism of sugars in this fruit. The variety 'Hass' is a popular cultivar that is grown extensively in South Africa. However this. cultivar has a tendency to produce two distinct fruit phenotypes: a normal sized variant and a small, undersized variant. Current literature suggests that the small fruit phenotype is characterised by an elevated abscisic acid (ABA) to cytokinin ratio and altered isoprenoid metabolism. The results presented in the current investigation represent the findings from a detailed study into the metabolism and transport of sugars in 'Hass' fruit in an attempt to characterise solute allocation in developing avocado fruit. Furthermore, the activities of sugar metabolising enzymes, routes of solute movement and polyphenolic contents of normal, small and ABA-treated fruit were compared and contrasted to evaluate the potential role of ABA in the induction and expression of the small fruit phenotype. The enzymes invertase, sucrose synthase (SSy) and sucrose phosphate synthase are involved in the metabolism of sucrose (Suc) and, hence, phloem unloading, post-sieve element transport and fruit growth. Although not the major sugar present, Suc was found in avocado phloem sap, and the enzymology for its metabolism was shown to exist in avocado fruit. It appears that sink strength is established during early fruit growth by high acid invertase activity, especially during the period of rapid cell division. As fruit growth progresses the activity of SSy and an enzyme responsible for the oxidation of perseitol (tentatively termed perseitol dehydrogenase) increases, suggesting that these enzymes play an important role in the supply of carbon during the linear phase of fruit growth. All Suc metabolising enzyme activity diminishes as the fruit approaches maturity. With the exception of SSy (in the cleavage direction), all enzymes assayed showed a general increase in relative rates of activity in small and ABA-treated fruit. Similarly, ABA-treatment of seed coat discs in vitro resulted in the elevation of insoluble and soluble acid invertase, SSy (in the synthesis direction), and sucrose phosphate synthase activity. Furthermore, both small and ABA-treated fruit were characterised by elevated total soluble sugars, glucose and fructose levels. These observations suggest that altered sugar metabolism, as a consequence of changes in endogenous ABA levels, may contribute to the occurrence of the small fruit. The seed coat represents an import link between the seed, the mesocarp and the parental plant tissues. Loss of seed coat and endosperm integrity accompanied fruit maturation and a reduction in the movement of solutes into the seed. An increase in polyphenolics in the seed coat tissue seemed critical in this reduced movement. Both the small and ABA-treated fruit were characterised by early senescence of the seed coat, which was accompanied by both a loss of transport into and out of the seed and premature maturation of the fruit. This premature seed coat senescence appeared similar to programmed cell death in tissues exposed to stress or elevated reactive oxygen species, stimuli that are often accompanied by elevated ABA levels. Callose was localised to the plasmodesmata and is proposed to play a role in the gating of, and hence movement through, these pores. Small fruit were characterised by a loss of symplastic continuity, as represented by fewer plasmodesmata, and reduced callose degradation. Comparison of callose content and rates of synthesis suggest that ABA-treatment, similarly, reduces callose catabolism. The association of ABA with both the premature senescence of the seed coat and a reduction in symplastic continuity, and, hence, a reduction in solute transport, further cements the potential role of ABA in the occurrence of the small fruit phenotype.
Some effects of drying rate and wet storage on aspects of the physiology and biochemistry of embryonic axes from diesiccation- sensitive seeds.
(2004) Ntuli, Tobias M.; Berjak, Patricia.; Pammenter, Norman William.; Smith, Michael Trevor.
Desiccation-sensitive seeds show differential viability characteristics during drying at different rates. A number of studies have demonstrated that rapid dehydration permits survival to lower water contents than does slower desiccation. The aim and objective of the present study was to test the hypothesis which states that rapid drying of desiccation-sensitive seeds removes water sufficiently fast to reduce the accumulation of metabolic damage. In addition, the hypothesis that wet storage subjects desiccation-sensitive seeds to mild, but increasingly severe, water stress causing oxidative damage if additional water is not supplied, was tested. In the present study, axes of germinating orthodox seeds of Pisum sativum and newlyshed recalcitrant counterparts of Quercus robur, Strychnos madagascariensis, Trichilia emetica, Trichilia dregeana and Avicennia marina were subjected to rapid or slow drying or wet storage. For those species where more than one harvest was investigated, differences were observed in water contents at shedding. For all the species studied, the dehydration rate could be described by an exponential and a modified inverse function for both desiccation regimes, and the water content remained constant with wet storage. The level of tetrazolium staining and germination percentage of axes decreased sharply drying and hydrated storage such that the marked decline took place at lower water contents upon rapid than slow dehydration. The conductivity of electrolyte leachate increased progressively during desiccation and moist storage of axes of all species investigated. Greater membrane leakage occurred upon slow, than rapid dehydration in axes of all species studied. Activities of respiratory enzymes which have a potentially regulatory role in glycolysis, phosphofructokinase (PFK), or the tricarboxylic acid cycle, malate dehydrogenase (MDH), and levels of the oxidized form of the coenzyme, nicotinamide adenine dinucleotide (NAD), of the enzymes of the electron transport chain, NADH dehydrogenases ofNADH-ubiquinone (coenzyme Q) reductase (complex I) and NADHcytochrome c reductase (complex IV), were monitored in the present investigation. v In addition, the role of free radical activity in the form of lipid peroxidation, which has been implicated in loss of viability in seeds, was examined by assaying the levels of hydroperoxides. The involvement of the free radical processing enzymes, superoxide dismutase (SOD), catalase (CAT) and glutathione reductase (GR), and the antioxidant, ascorbic acid (AsA), was also ascertained. The activity of PFK in axes of P. sativum remained constant during drying and wet storage. However, PFK activity increased as rapid dehydration and hydrated storage of Q. robur axes proceeded. In contrast, the activity of PFK in axes of Q. robur decreased during slow desiccation. Similarly, PFK activity was reduced upon drying, and moist storage, of T. dregeana axes such that higher activity of PFK was seen during rapid than slow dehydration. The activity ofPFK inA. marina axes also declined upon desiccation. The activity ofMDH in axes of P. sativum was also unchanged during drying and wet storage. However, an increase in MDH activity was recorded in Q. robur axes during dehydration and hydrated storage such that the activity of MDH was higher upon slow than rapid desiccation. In contrast, MDH activity in axes of T. dregeana decreased as drying proceeded. Similarly, the activity of J\.1DH declined during dehydration and moist storage of A. marina axes. An increase in the level of NAD occurred in axes of P. sativum during drying. In contrast, a decrease in NAD levels was seen upon dehydration and wet storage of Q. robur axes such that the level of NAD was higher upon rapid than slow desiccation. There was an enhancement of the level of NAD in axes of T. dregeana during hydrated storage. Conversely, NAD levels declined during drying ofA. marina axes. A decrease in the level of hydroperoxides in axes of P. sativum was seen as rapid drying proceeded. In contrast, hydroperoxide levels increased during wet storage of P. sativum axes. Similarly, the levels of hydroperoxides were enhanced upon dehydration and hydrated storage of Q. robur axes such that they were higher in axes during slow desiccation compared to those dried rapidly. Conversely, the hydroperoxide level in axes of T. dregeana was reduced upon rapid dehydration. In contrast, an elevation of the level of hydroperoxides was observed during moist storage. The levels of hydroperoxides remained constant as desiccation and wet storage ofA. marina axes proceeded. vi The activity of SOD in axes of P. sativum decreased during rapid drying. In contrast, SOD activity increased upon slow dehydration and wet storage ofP. sativum axes. There was a decline in the activity of SOD in Q. robur axes during slow desiccation. Similarly, SOD activity was diminished upon drying of axes of T. dregeana. The activity ofSOD in T. dregeana axes was enhanced during hydrated storage. An elevation in SOD activity also took place during rapid dehydration and moist storage of axes ofA. marina. The activity of CAT did not change during drying of axes of P. sativum. However, a decrease in CAT activity in Q. robur axes was seen upon slow dehydration and wet storage. Similarly, the activity of CAT declined as desiccation of axes of T. dregeana proceeded. In contrast, CAT activity inA. marina axes increased during slow drying. Whereas the activity of GR in axes of P. sativum increased during drying and wet storage, GR activity decreased in A. marina axes upon all treatments such that the activity ofGR was higher during rapid than slow dehydration. GR activity also declined upon slow desiccation and hydrated storage ofaxes of Q. robur. Similarly, the activity of GR in T. dregeana axes was reduced during moist storage. Finally, a decrease in the level of AsA in axes of P. sativum took place during drying. Nonetheless, dehydration and wet storage of Q. robur axes were associated with no siginificant change in AsA levels. There was also a decline in the level of AsA in axes of T. dregeana as rapid desiccation proceeded. Similarly, a reduction in AsA level occurred upon slow drying ofaxes ofA. marina. The results presented here are consistent with the observation that drying and wet storage adversely affected the respiratory enzymes, PFK, MDH and NADH dehydrogenase. It is suggested that the resultant metabolic imbalance led to more leakage of electrons from the mitochondrial electron transport chain than normal, and through lipid peroxidation increased levels of hydroperoxides. In addition, dehydration and hydrated storage may depress the activities of free radical processing enzymes, SOD, CAT and GR and levels of antioxidant, AsA. This phenomenon was less pronounced during rapid, in comparison to slow, desiccation and moist storage. However, it appears that the above biochemical events are overtaken by physical damage at higher water contents in the highly recalcitrant seeds. It was concluded that the differential effects of VII the drying rate and wet storage on responses of desiccation-sensitive seeds varies with tissue, harvest, species and the degree of desiccation sensitivity.
A systematic study of Leonotis (Pers.) R. Br. (Lamiaceae) in southern Africa.
(1995) Vos, Wayne Thomas.; Edwards, Trevor John.; Van Staden, Johannes.; Smith, Michael Trevor.
The southern African species of Leonotis (Pers.) R. Br. are revised. Nine taxa are recognised including a new species and two new subspecies. A multidisciplinary approach was adopted involving: floral, vegetative, seedling and fruit morphology, karyology, palynology, pollination biology, phytogeography, phytochemistry and cladistics. Of these characters the most important in delimiting species is leaf morphology. Phytochemistry revealed qualitative and quantitative differences in essential oils between taxa. Germacrene is the dominant essential oil in seven species, particularly in L. dubia E. Mey. and L. nepetifolia (L.) R. Br. Caryophyllene dominates in L. randii S. Moore. Although certain species have characteristic aromas, no inter-specific differences are apparent in the levels of α-copaene, the dominant volatile essential oil extracted with Tenax. Essential oil data is in partial agreement with the phylogeny and species concepts presented in this document. Omithophily accounts for most fruit set in perennial species but in the annual species (L. nepetifolia) autogamy prevails. Neither omithophily nor entomophily improved nutlet vigour which was gauged from nutlet germination. Leonotis species are predominantly pollinated by a variety of sunbirds, although bees are also involved. Nectar is sucrose-dominant in perennial species but hexose-dominant in the annual. The majority of species are concentrated along the eastern seaboard with the eastern Cape forming the centre of diversity. The widespread distribution and tropical affinities of L. leonurus (L.) R. Br., suggest an origin for the genus in tropical Africa. Migration to southern Africa and subsequent speciation are suggested. Habitat diversity and edaphic conditions probably played an important role in the evolution of narrowly distributed xerophytic species. A cladistic analysis re-emphasised that Leonotis is a monophyletic group of closely related species. The lack of floral divergence in perennial species indicates the constraints imposed by ornithophily. The annual, L. nepetifolia, is highly derived and displays numerous autapomorphies. The morphology, distribution and tropical affinities of L. leonurus are interpreted as pleisiomorphic. The proposed phylogeny is reasonably compatible with phytogeographic data.