VUT DigiResearch
Digiresearch is the VUT electronic open access archive. Its purpose is to collect, distribute and preserve the research performed by the VUT Research community
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Recent Submissions
An investigation of the immunomodulatory effects of crude extracts from Carpobrotus edulis on macrophages in vitro
(Vaal University of Technology, 2024-12) Laloo, Neelan; Ssemakalu, S. S., Prof.; Pillay, M., Prof.
Carpobrotus edulis is an edible halophyte which is used widely in traditional medicine in South Africa for a variety of ailments. The plant has antioxidant, anti-inflammatory, antibacterial, antifungal and anticancer properties and is used in the treatment of skin and wound infections, insect bites and mouth and throat infections. Despite its wide use as a medicinal plant, empirical research focusing specifically on its immunomodulatory effects remains sparse. This study aimed to bridge this knowledge gap by rigorously investigating the immunomodulatory effects of C. edulis on RAW264.7 macrophage cells using aqueous and methanol extracts. Two 2³ factorial designs were used to determine the optimal parameters to maximize the yield of crude leaf extracts using the maceration technique. The variables included solvent (water or methanol), pH (5 or 9), extraction temperature (25 or 40℃) and extraction duration (72 or 168 h). Design Expert 11 software was used to determine which extraction parameters produced the highest yield and phytochemical content. Quantitative phytochemical analysis was conducted using the Folin-Ciocalteau and Aluminium Chloride methods to determine total phenolic concentration (TPC) and total flavonoid concentration (TFC), respectively. The highest extraction yield (31.03 ± 0.58%) with water obtained at pH 9, for 72 hours at 40℃, had a TPC of 6.42 ± 0.03 mg Gallic Acid Equivalent per gram of extract. The highest yield with methanol (64.21 ± 2.12%) was obtained at pH 9, 168 hours and 40℃. The highest TPC (7.44 ± 0.50 mg GAE/g of extract) with methanol was obtained at pH 9, 72 hours of extraction, and 25℃. The radical scavenging activity of the extracts was determined using the ABTS and DPPH assays. For the aqueous extracts, the conditions which produced the highest yield and TPC (pH 9, 72 h and 40℃) also produced the lowest IC50 values for both assays – 140.77 μg/mL for ABTS and 298.28 μg/mL for DPPH. However, with methanol different extraction parameters produced differing results. The lowest IC50 value for the ABTS assay (26.79 μg/mL) was obtained at pH 5, 168 hours of extraction and 25℃; whereas the lowest IC50 value for the DPPH assay (109.84 μg/mL) was from the extract obtained at pH 9, 72 hours extraction time and at 25℃. All the IC50 values were higher than those for the positive control, ascorbic acid. Strong correlation was found between the TPC and radical scavenging activity for the aqueous and methanol extracts – as the TPC increased, the IC50 decreased. Using the DPPH results, r values were -0.952 and -0.759 for the aqueous and methanol extracts, respectively. For the ABTS assay, the correlation coefficients were -0.879 and -0.320 for the aqueous and methanol extracts, respectively. The effect of the extracts on the metabolic activity and protein content of the RAW264.7 cells was determined using the CellTiter Blue® (CTB) and Sulforhodamine B (SRB) assays, respectively. The cells were metabolically active even after treatment with 3000 μg/mL of the aqueous extract. However, protein content decreased as the aqueous extract concentration exceeded 1000 μg/mL. The aqueous extract with the highest metabolic activity, using the CTB assay, was obtained at pH 5, 168 hours and 40℃. However, the SRB assay showed the highest protein content in an extract obtained using the same time and temperature, but at pH 9. RAW264.7 cells treated with the methanol extracts showed increases in metabolic activity of greater than 1200% than the untreated control cells. These extracts were both obtained at 40℃ and after 72 hours of extraction except that one was at pH 5 and the other at pH 9. The latter parameters also produced the highest protein content. The potential pro- or anti-inflammatory effect of the extracts was determined using the Griess assay for nitric oxide production. RAW264.7 cells stimulated with lipopolysaccharide (LPS) at 100 ng/mL were treated with extract at concentrations from 1 μg/mL to 1500 μg/mL. In the cells treated with three of the eight aqueous extracts, the nitric oxide production increased as the extract concentration increased. Cells treated with an aqueous extract obtained at pH 9, and 72 hours of extraction at 25℃. produced the highest NO. Two treatments did not produce NO for concentrations of the aqueous extract above 10 μg/mL; these extracts were produced at 40℃ and 168 hours at either pH value. None of the cells treated with methanol extracts (at concentrations above 100 μg/mL) produced NO. The aqueous extract obtained at pH 9, 40℃ and 168 hours and the one for methanol obtained at pH 9, 40℃ and 72 h were selected for further study based on their viability, proliferative potential, and low nitric oxide production. Different concentrations of these were used to treat LPS-stimulated RAW264.7 cells to obtain RNA at 24 and 48 hours. The RNA was converted to cDNA, which was amplified using qPCR to determine the expression levels of three genes associated with the M1 macrophage phenotype (Stat1, NF-κB, Nos2) and three associated with the M2 phenotype (Stat6, Irf4, Arg1). The aqueous extract did not trigger an inflammatory response, as the expression levels (calculated as fold change 2-(ΔΔCt)) were close to zero for NF-κB and Nos2 suggesting that the expression was significantly downregulated (p = 0.0009). The aqueous extract at 1500 μg/mL did significantly (p = 0.0002) increase in the expression of Stat1 after 24 hours. The aqueous extract did not increase the expression of the M2 genes either. Only a slight increase in Irf4 expression was observed in comparison with the untreated control cells (p > 0.05). The response of the M1 genes to the methanol and aqueous extract was similar. A significant (p < 0.0001) increase in Stat6 and a slight increase in Arg1 (p = 0.0179) were detected after 24 hours in response to the methanol extract at 250 μg/mL. To further investigate the response of the cells to the two C. edulis extracts, a pre-mixed Luminex MAP® cytokine kit was used to detect the expression of cytokines and chemokines related to the M1 and M2 macrophages. At concentrations up to 1000 μg/mL and 48 hours of treatment, the aqueous extract significantly increased the expression of the M1 molecules CCL2 and GM-CSF (p < 0.0001) and at up to 500 μg/mL, CXCL10 (p = 0.0003). Molecules associated with the M2 phenotype only increased in concentration after 48 hours at extract concentrations up to 1000 μg/mL. TNF-α and IL-6, which are expressed by both M1 and M2 macrophages increased in concentration after treatment with the aqueous extract up to 1000 μg/mL (p < 0.0001 for both). A similar response was observed for cells treated with the methanol extract up to 100 μg/mL, but onlythe IL-6 concentration was significantly different from the control (p < 0.0001). The M1 molecules CCL2, GM-CSF and CXCL10 increased significantly in comparison with the untreated cells (p < 0.0001) when treated with up to 100 μg/mL of the methanol extract. These results suggest that in the absence of a strong anti-inflammatory response, the cells are being polarized towards an M1 phenotype. High concentrations of GM-CSF attract monocytes and macrophages to the site of injury. This suggests that the two C. edulis extracts used here are nontoxic and have an immunostimulatory effect on the RAW264.7 cells. This study showed that the therapeutic effects of C. edulis appear to be dependent on extraction conditions and the concentration of the extracts.
The effect of carpobrotus edulis and cotyledon orbiculata on the migration, proliferation and differentiation of keratinocytes in vitro
(Vaal University of Technology, 2020) Terblanche, Unisa; Ssemakalu, Dr. C.C.; Pillay, Prof. M.
Cutaneous wounds are an epidemic that affects millions of people worldwide. Healing and regeneration of wounds is a complex process that involves several molecular and cellular pathways. Traditional healers have used plants for the healing and regeneration of cutaneous wounds for centuries. In South Africa, two species, Carpobrotus edulis and Cotyledon orbiculata, have been used in traditional medicine for the treatment of various skin conditions and wounds. There is limited empirical evidence on the effectiveness of some of these commonly used plants on wound healing. The aim of this study was to use a factorial design to develop a protocol for optimal extraction of bioactive compounds from Carpobrotus edulis and Cotyledon orbiculata using water and methanol as extractants and evaluate the effectiveness of these extracts on the migration, proliferation and differentiation of keratinocytes in vitro. To achieve this the study followed a 2 step integrated approach. In the first part of this study, 23 full factorial designs were employed to investigate the effect of different extraction parameters on the yield of crude extracts obtained from the leaves of Carpobrotus edulis and Cotyledon orbiculata, respectively. The impact of these parameters on the phytochemical constituents were assessed using standard colorimetric tests, whereas the Folin-Ciocalteau and Aluminium Chloride colorimetric assays were used to determine the impact on the total phenolic (TPC) and total flavonoid (TFC) contents, respectively.
This was followed by the utilisation of two cell viability assays, CellTiter® Blue (CTB) and Crystal violet (CVS) staining, to assess the effect of these rude extracts on an immortalised human skin keratinocyte (HaCaT) cell line, in vitro. The response values from these assays were used in a 25 factorial design to develop the best-fit regression model to predict which aqueous and methanolic extract (for each plant) expressed the highest proliferative potential. A pH of 9 for 72 h at 40°C proved to be the best extraction conditions for the aqueous extraction of Carpobrotus edulis, resulting in a yield of 31.03% whereas the methanolic extract produced a maximum yield of 64.21% at a pH of 9 for 168 h at 40°C. For Cotyledon orbiculata, the highest yield of 44.13% was achieved for the aqueous extract at a pH of 9 for 4 h at 60°C whereas 50% methanol for 168 h at 25°C resulted in a maximum yield of 33.64% for the methanolic extraction. Preliminary tests of the plant leaf extracts indicated the presence of reducing sugars, phenols, tannins, flavonoids, cardiac glycosides, diterpenes, phytosteriods and triterpenoids in both species. The highest TPC and TFC for the aqueous extracts of Carpobrotus edulis were 16.43 ± 0.07 mg GAE/g and 7.42 ± 0.53 mg QE/g respectively, whereas values of 18.92 ± 0.23 mg GAE/g and 6.88 ± 0.44 mg QE/g were obtained for the methanolic extracts. The highest TPC and TFC for the aqueous extract of Cotyledon orbiculata were 38.79 ± 1.07 mg GAE/g and 2.60 ± 0.04 mg QE/g, respectively. For the methanolic extract, maximum values for TCP and TFC were 56.49 ± 1.79 mg GAE/g and 12.35 ± 0.21 mg QE/g, respectively. The aqueous extract of Carpobrotus edulis increased the viability of the HaCaT cells by approximately 67% ± 3.93%, whereas a similar extract of Cotyledon orbiculata only increased cell viability of HaCaT by 29% ± 0.99%. Based on the experimental observations and developed regression models, the extraction parameters that predicted the highest proliferative potential across both assays were a pH of 5 for 168 h at 40°C for the aqueous extract of Carpobrotus edulis, and a pH of 5 for 168 h at 25°C for the methanolic extract. For the aqueous extract of Cotyledon orbiculata, a pH of 7 for 4 h at 30°C predicted the highest proliferative potential whereas best results for the methanolic extract was predicted to be achieved with full strength methanol, for 168 h at 25°C. In the second part of this study, these optimised extraction models were used to assess the in vitro wound healing ability of Carpobrotus edulis and Cotyledon orbiculata as a measure of its ability to promote re-epithelialisation. Cytotoxicity of the extracts that was assessed morphologically and with the Lactate Dehydrogenase (LDH) Cytotoxicity assay showed that the aqueous extract of Carpobrotus edulis showed little to no morphological changes at concentrations ranging from 100 to 1000 μg/mL which correlated with the non-significant (p > 0.05) LDH activity whereas the methanolic extract exhibited morphological changes at all tested concentrations. On the contrary, both the aqueous and methanolic extracts of Cotyledon orbiculata induced morphological changes of the HaCaT cells at most concentrations. The significant (p < 0.05) increase in LDH activity for the aqueous (8.4% ± 2.54% to 26.9% ± 2.38%) and methanolic extract (5.3% ± 1.14% to 10.6% ± 1.64%) verified this result. The proliferation of HaCaT cells after treatment with the crude extracts, measured with the sulforhodamine B (SRB) assay and the xCELLigence Real-Time Cell Analysis (RTCA) system, showed a significant reduction in doubling times of the cells when they were treated with the aqueous extract of Carpobrotus edulis. The methanolic extracts of the species showed some increase in proliferation rates with 500 to 1000 μg/mL after 24 h of treatment, but this was not supported with the RTCA system. For the Cotyledon orbiculata, the SRB assay showed that the aqueous extract at concentrations of 1, 10 and 25 μg/mL significantly increased the proliferation of HaCaT cells after 48 h of treatment; however, the RTCA system did not support this proliferation rates or doubling times. For the methanolic extract, an antiproliferative effect was observed for all treatments 24 h after exposure. This was supported by the absence of any evidence of a proliferative effect or decreased doubling time using the RTCA system. The migration potential of HaCaT cells, evaluated using an in vitro scratch assay and the RTCA system, showed that the aqueous extract of Carpobrotus edulis increased the wound closure by 31% compared to the untreated cells between concentrations of 1000 to 2000 μg/mL. The methanolic extract increased the migration rate by 17% only at 10 μg/mL. On the contrary, the aqueous extracts of Cotyledon orbiculata decreased the rate of migration by 10% and 30% for both the 10 μg/mL and 50 μg/mL concentrations, respectively. The methanolic extract also decreased migration rates for the 10 μg/mL and 50 μg/mL by 38% and 37%, respectively. The expression of selected cytokines by migrating HaCaT cells were assessed using a Luminex MAP® cytokine kit. Cytokine expression for Carpobrotus edulis showed that both solvent extracts did not significantly influence the expression of TNF-α, IL-6, and IL-8. It was, however, found that the expression of TGF-α and VEGF were reduced for both solvent media. Cytokine expression was not investigated for Cotyledon orbiculata as the plant showed an inhibitory effect on migration for both aqueous and methanolic extracts. The evaluation of the effect of the extracts on keratinocyte differentiation, measured by the expression of differentiation markers K1,10 and involucrin showed either no change, or a reduction in K1,10 expression for both aqueous and methanolic extracts for both species. Involucrin expression was inconclusive for both plants. The results of this study showed that within a concentration range of 1000 and 2000 μg/mL, the aqueous extract of Carpobrotus edulis promoted keratinocyte proliferation and migration. This supports the ethnomedicinal use of the plant for the treatment of cutaneous wounds. While Cotyledon orbiculata is also used as folk medicine for wound healing, this study found no evidence to support such a claim.
Employees' perceptions of the effects of retrenchment on job stress and organisational commitment in a mining company
(Vaal University of Technology, 2016-11) Seteni, Landiswa Pilvia; Joubert, P. A., Dr.; Dhurup, M., Prof.
In the decade before 2005, South African organisations have had to cope with an ever-increasing rate of local and global changes. There have been considerable and ongoing socio-political changes, resulting from new Government regulations. Most organisations have experienced some type of downturn, whether due to external business factors or poor internal performance. A typical response to organisational decline is retrenchment. Retrenchment is attributed to cyclical downturns, market losses or other economic factors, which oblige the employer to reduce the labour force numbers.
Though there are so many ways of responding to organisational failure, this study focused only on retrenchment. A number of psychologists and human sciences researchers have studied the results of job loss due to retrenchment. Retrenchment brings loss of skills, loss of morale and commitment, as well as physical and mental health degradation, including stress that results in employees withdrawing physically and emotionally.
The main purpose of this study was to outline the employees’ perceptions of the effects of retrenchment on job stress and organisational commitment in a mining company. The research methodology used in this study is a combination of a literature review and an empirical study. The probability sampling technique, which entails using simple random sampling, was used to select the sample of the study. The primary data were collected using a questionnaire. The measuring instrument contained 43 items. The measuring instrument was pilot-tested with 50 respondents three weeks before the main survey; the questionnaire was self-administered to the participants. For the main survey, data from 294 respondents were collected and analysed. Participants in the study involved surface employees, including management, administrators, engineers and artisans.
Data were analysed with the Statistical Package for Social Sciences (SPSS) version 23.0. The Cronbach’s alpha coefficients of the various sections of the measuring instrument were computed to establish construct validity. Content validity was ascertained by pre-testing the questionnaire with the employees in the mine in question. Correlations were used to establish convergent validity of the constructs. Convergent validity was assessed to measure the degree of linear association of variables using Pearson’s correlation coefficient. Discriminant validity was achieved by using regression analysis to show items loaded onto different factors in various sections which had more than one factor.
Analysis was done using descriptive statistics on the demographic information of respondents. The results were also interpreted through correlation and regression analysis. The results showed that the effects of retrenchment did not have a negative impact on job stress (time stress and anxiety). This may be due to the mine in question planning and consulting with the employees prior to the retrenchment process. The results also showed that job stress (time stress and anxiety) is negatively associated with organisational commitment. Subsequent to these findings, it is recommended that employees (survivors) be updated on their future in the company in question. This could be done through regular workshops and counselling of employees.
In order to motivate and engage employees, and thereby contribute to employee commitment, it was also recommended that organisations create open, supportive and fair organisational and team cultures, and ensure jobs are clearly aligned with organisational goals and have appropriate levels of autonomy, support and career development. Given that job satisfaction plays a huge role in organisational commitment, it was further recommended that managers need to actively improve their organisation’s job satisfaction to employees in order to achieve a higher level of organisational commitment.
The study concludes by recommending that communication between employees and management should take place on a regular basis which can serve as a fundamental tool to retrenchment in the following manner:
• Survivors will feel more committed to the organisation when the basis of retrenchment addresses the circumstances in the external environment, rather than the enrichment of shareholders or top management.
• Communicating a clear vision of how retrenchment will benefit stakeholders, increases commitment since survivors can see a clear future for the organisation.
• When survivors are treated with dignity and respect, they will feel more committed to the organisation because they feel appreciated.
Determination and improvement of wear and abrasion properties of Glass Reinforced Plastic (GRP) piping for slurry applications
(Vaal University of Technology, 2014-09) Van der Schyff, A.; Mendonidis, P., Prof.
Glass Reinforced Plastics (GRP) are composite materials comprising a resin matrix reinforced with glass fibers, and are well known for their excellent corrosion resistance and high strength-to-weight ratios. However, their use in slurry transporting pipes is uncertain because the wear resistance of GRP pipe materials is not well documented due to the difficulty encountered in standardizing mechanical properties of GRP pipes. The aim of this project was to identify the main variables that influence the wear and abrasion properties of GRP pipes and the differences of abrasion properties between the continuous and discontinuous filament wound pipes that are manufactured by Amitech SA. In order to achieve the aim of the project, the mechanisms of wear in ductile and brittle matrix GRP composites were investigated. The Pin-on-disk and Jet-impact tests were identified as the best abrasion tests suited for testing GRP composites. Data collected from these tests indicated that the resin type, curing, additive content and- type, fiber content and manufacturing methods are factors affecting wear resistance. It was further found that hard surfaces provided better resistance to abrasive wear, whereas ductile matrices performed better under erosive wear.
Influence of job embeddedness on job satisfaction and job performance among employees within South African Police Service Academies
(Vaal University of Technology, 2022-05) Tala, Ratsela Johannes; Marais, Christel, Dr.; Grobler, Bennie, Prof.
Background: The South African Police Service (SAPS) faces various challenges relating to service delivery, which is viewed as stressful, demanding, and dangerous. Members work long, stressful hours under adverse and challenging working conditions. These demanding working conditions correlate with experienced job embeddedness, job satisfaction and job performance.
Moreover, in addition to these challenges police employees suffer from physical ailments, such as heart condition, hypertension and sugar diabetes, mellitus, and psychological problems, such as cynicism: feelings of hopelessness, suicidal thoughts and suicide. These challenges may lead to absenteeism, resignations and ineffective organisational performance.
Many SAPS personnel at various academies have been employed in the Human Resource Development (HRD) for a prolonged time, and employee turnover is low. This suggests that they are committed to their job and experience job embeddedness. Research has shown that job embeddedness might predict job satisfaction and performance. The study aimed to investigate this relationship in the context of SAPS academy employees to inform SAPS management in assisting police officers in performing their role despite the challenges they face.
Training is a high priority in the SAPS, the academies of which are expected to prepare future police officers to fill their roles effectively. However, the SAPS academies, which are expected to provide training and development, have limited human, financial and logistical resources owing to current political and socio-economic circumstances. Therefore, although employees in these academies may face challenges to their job embeddedness, job satisfaction and job performance, they remain in their jobs, which is a situation that the study aimed to explore.
Aim: The study described in this thesis aimed to investigate the influence of job embeddedness on job satisfaction and job performance amongst employees within South African Police Service academies, viewing it as a holistic construct.
Methodology: The research methodology followed to conduct the study was a combination of a literature review and an empirical study. Existing scholarship informed a theoretical understanding of the topic. An empirical investigation was conducted to achieve the aim mentioned above. In addition, a quantitative descriptive research design was adopted to investigate a sample of 400 employees working in the 23 academies nationally. Data were collected using a structured questionnaire as the collection instrument which was piloted with 25 respondents and then hand-delivered to the respondents of the main study. Simple random sampling accurately reflected the larger population (N=956).
Results and Discussion: The research results indicated that job performance and job satisfaction are causally related via job embeddedness acting as a mediator. Moreover, the relationship between job performance and job satisfaction was found to be non-recursive. The total effect of job performance on job satisfaction is larger (.5956) than the total reciprocal effect between job satisfaction as a predictor and job performance as an outcome (0.4061). The research supported the view that job satisfaction and job performance have a reciprocal relationship moderated by other variables such as job embeddedness. This finding does not support the initial hypothesis that job satisfaction acts as a mediator between job embeddedness as a predictor and job performance as an outcome.
General: As little research has been conducted on the relationship between job embeddedness, job satisfaction and job performance in the SAPS and the public sector in South Africa, relevant literature was not readily available for the study. In addition, the study had other limitations, which are highlighted in this thesis to inform similar future studies. However, the results provided insights that researchers in South Africa might find helpful, although the effect of job embeddedness as a holistic construct on various variables needs to further investigated in the context of the SAPS.
Conclusions: The study revealed that job performance, directly and indirectly, affects job satisfaction with job embeddedness acting as a mediating variable. The association between job satisfaction, job performance and job embeddedness as dependent variables and gender as an independent variable found no significant relationship. However, the relationship between race and first language as independent variables and job performance and job satisfaction as dependent variables differed significantly.
In the questionnaire, white respondents agreed significantly less strongly with the job satisfaction and job performance constructs than the black African and coloured respondents. This difference was present in the task and contextual performance sub-dimensions of job performance. The Nguni first language group agreed significantly more strongly with the job performance and job satisfaction constructs than the Afrikaans first language group. This result was also true for the two sub-dimensions of job performance, namely task and contextual performance, with task performance being perceived as slightly more important than the contextual performance by the Nguni group. No significant associations could be found between age, educational level, marital status and the number of years employed as a government employee as independent variables and the three dependent variables of job embeddedness, job satisfaction and job performance.
Recommendations: The study led to recommendations for management practice, which might be helpful to the management of the human resource development division of the SAPS. The study provided essential information about the importance of job embeddedness in the working environment and its effect on employees’ job performance and job satisfaction who are expected to provide quality training in the SAPS. In addition, the study led to recommendations for future research by enhancing the knowledge base related to job embeddedness in the context of the South African public service sector. In conclusion, job embeddedness as a holistic construct and its relationship with other variables, needs to be explored further, especially in the context of the SAPS and its academies.