Theses (MSc. Medical and Health Sciences)

BACKGROUND: Informal caregivers are friends or relative who offer to care for stroke patients after discharge. The care deliveries typically occur without formal training or compensation. The COVID-19 pandemic may increase the level of responsibility of caregivers, resulting in potential negative effects on their overall well-being. METHODS: A cross-sectional study of 94 unpaid informal caregivers of stroke survivors completed an online survey. The stroke caregivers’ burden was measured using the Caregiver Burden Inventory (CBI). Social connection such as social isolation, loneliness and perceived social support was measured using the Lubben Social Network Scale-6 items, UCLA-3 items and Duke Social Support Index (DSSI-11 item), respectively. Sociodemographic data were also collected and analysed. RESULTS: Multiple linear regression analysis revealed that social support (β = -0.34, t = -3.46, p < .001) and loneliness (β = 0.33, t = 3.35, p = .001) significantly predicted the stroke caregivers’ burden. Social support had the major predictive effect, explaining 12% of the variance in caregivers’ burden (R2 = 0.12). Loneliness explained 11% (R2 = 0.11) and 8% (R2 = 0.08) of the variance in developmental and physical burdens, respectively. Social isolation was excluded and not a significant predictor of caregivers’ burden. Caregivers with tertiary education reported significantly higher burden (β = 0.28, t = 2.81, p = .006) than caregivers with secondary education. Education was also a significant predictor of developmental burden (β = 0.25, t = 2.47, p = .016), physical burden (β = 0.29, t = 2.89, p = .005), and emotional burden (β = 0.22, t = 2.12, p = .037). CONCLUSION: The burden experienced by stroke caregivers is linked to their social support, loneliness and educational levels. These findings provide further insight on the role of social support during COVID-19 and its contribution towards caregiver’ burden in an unprecedented time. Interventions that aim to enhance social support for caregivers could help to mitigate this issue and improve their overall well-being.

Dietary fat when consumed reduces hunger and impairs food intake by eliciting satiety signals and these signals are evoked by entry of triacylglycerol after hydrolization to fatty acids into the small intestine. 1,3-dipalmitoyl-2-oleoyglycerol (POP-), 1,3-distearoyl-2-oleoylglycerol (SOS-) and 1,2,3-triolein (OOO-) type of fats have different melting characteristics that may affect postprandial blood lipids, gut hormone concentrations, insulinemic response and selected cardiovascular disease markers in human volunteers. The main objective of this study is to compare the effects of edible fats with either palmitic acid (16:0) (palm mid-fraction) or stearic acid (18:0) (shea stearin) predominantly at the sn-1 and sn-3 positions on postprandial lipemia and gut hormone concentrations. A randomized, double-blind crossover (3 × 3 arms) orthogonal Latin-square design was used on 36 healthy adults (18 males, 18 females; mean age = 23 years). Each subject received 3 different test muffins (each containing 53 g of test fat) in random order separated by 2 weeks over a 6-week period. The test fats of different melting points were palm mid- fraction (PMF; POP-rich), shea stearin (SS; SOS-rich) and high- oleic sunflower oil (HOSF; OOO-rich) During a postprandial test, each subject was provided with a test muffin plus milkshake (total 3.67 MJ or 876 kcal) in the morning and blood samples were collected at half-hourly intervals until 4.0 hours. No significant difference (p>0.05) was observed between the 3 test meals for postprandial responses in plasma TC, Lp(a), apo(B), NEFA, GLP-1, PYY, ghrelin, VAS, PAI-1, IL-6, TNF-α, glucose, insulin and satiety (VAS scores). Plasma TAG peaked at about 4 hours; levels in the PMF- and HOSF- subjects were significantly higher (p<0.05) compared with SS-subjects after 90 minutes. PMF and HOSF exerted a higher postprandial GIP response (p<0.05) as compared to SS. Plasma C-peptide levels, as a measure of insulinemic response, rose sharply 5.5- folds in all groups, peaking after 90 minutes; levels in the SS group declined at a faster rate (p<0.05) than in the PMF- and HOSF- groups. The POP- and OOO- fats induced similar effects on all the biochemical/physiological outcome measures investigated. In contrast, the SOS- type fat (shea stearin) induced a slower rise (p<0.05) in postprandial TAG and GIP levels and a faster return of plasma C-peptide levels to baseline.

Airborne algae and cyanobacteria (AAC) have received much attention as their occurrence have been documented in several studies, including indoor environment. There has been concern whether AAC can trigger allergic response and thus, they may have ill health effects, especially in relation to sick building syndrome (SBS). There have been very few studies reported on the allergenicity of AAC. The main objective of this study was to assess the allergenic potential of three AAC, namely Scenedesmus sp., Cylindrospermum sp. and Hapalosiphon sp.1, which were isolated from an office building in Kuala Lumpur. The study was based on a mouse model, with the suspension of AAC powder in phosphate buffered saline (PBS) administered via nasal route. Naïve mice were sensitised to 1 mg/mL of AAC for 10 consecutive days during Phase I immunisation. In Phase II immunisation which was given after five resting weeks, the mice were re-challenged with 20 μL of AAC at two concentrations (0.01 and 1 mg/mL) for three consecutive days before they were sacrificed. The immune response was assessed based on white blood cell profiles, quantification of humoural and intracellular expression of IgE, TH2 predominant and inflammatory cytokines and histopathological examination. In general, the three AAC elicited greater immune response in mice challenged with high (1 mg/mL) concentration of AAC than with low (0.01 mg/mL) concentration. In mice challenged with Scenedesmus sp. at both low and high concentrations, there were elevated peripheral eosinophil and monocyte counts. Significantly higher levels of serum IgE (P<0.05) and IFN-γ (P<0.05) compared to negative control were also observed. The data collected 48 h after the last dose of immunisation also indicated possible cause of allergic airway inflammation, a chronic manifestation of respiratory allergy upon nasal exposure to the algal particles. On the other hand, Cylindrospermum sp. and Hapalosiphon sp.1 did not elicit type-1 hypersensitivity as both eosinophil counts and serum IgE levels were not elevated in the sensitised mice. The results suggest that the two airborne cyanobacteria provoked immune response differently. In mice challenged with Cylindrospermum sp. (1 mg/mL), there were elevated serum IL-4 (22.1 pg/mL, P<0.05). However, the increase in peripheral neutrophil counts (9.2%) against Cylindrospermum sp. (1 mg/mL) was not significant when negative control was compared. In comparison, in mice challenged with Hapalosiphon sp.1, there were elevations of serum IL-6 (4.8 pg/mL, P<0.05) yet the observed increase of peripheral lymphocyte counts (9.2%) was not statistically significant. In addition, the elevations in serum IFN-γ levels were highest in mice nasal challenged with Cylindrospermum sp. (71.9 pg/mL, P<0.05), followed by Hapalosiphon sp.1 (50.6 pg/mL, P<0.05). Taken together, the results suggest that nasal exposure to Cylindrospermum sp. and Hapalosiphon sp.1 may provoke inflammatory immune responses in the airway. Further studies are warranted to assess the potential health impact due to the allergenic potential of the AAC, especially with regards to the effect of continual exposure to low concentration of the algal particles over extended period of time.

Diabetes mellitus is a chronic metabolic disease and predicted to increase about 366 million cases globally by end of 2030. Type 2 Diabetes is characterized by abnormal insulin secretion followed by inability of beta cells to compensate for insulin resistance. Most individuals diagnosed with diabetes are found to be obese and obesity as well is one of the major risk factors contributing to diabetes. Although modern medicine is used to control blood glucose level in diabetic patients but there is demand for natural products as anti-diabetic agents due to the side effects seen in the current drugs. In this study, we explore Nephelium lappaceum.L’s (or locally known as rambutan rind) ability as an anti-hyperglycemic agent in a type 2 diabetic animal model. The rind of N. lappaceum is found to consist of high antioxidant activity and possess in-vitro anti-hyperglycemic activity. This study is aimed initially to prepare standardised ethanolic extracts of N. lappaceum rind, evaluate toxicity effects in rat model, develop obesity induced type 2 diabetes rat model and finally investigate the anti-glycemic effects of Nephelium lappaceum rind in the diabetic rat model. Ethanolic extraction was followed as previously described to obtain N. lappaceum rind extract. Geraniin being the major bioactive compound in N. lappaceum rind was detected in LC/MS and quantified using HPLC chromatogram method to ensure a standardised sample was prepared. Biological standardisation was also carried out by subjecting the N. lappaceum rind to both alpha glucosidase and alpha amylase assay’s and to establish its in-vitro anti-hyperglycemic activity. Toxicity studies of N. lappaceum rind were carried out in Sprague dawley rats. In the acute toxicity study, rats were given 50mg, 200mg, 1000mg and 2000mg/kg of N.lappaceum rind orally for 14 days whereas in the sub-chronic toxicity study the rats were given a low and high concentration of N. lappaceum rind for 28days (500mg and 2000mg/kg). Type 2 diabetes was seen to develop in the rat model by feeding them with a high fat diet for 12 weeks and injected with 55mg/kg streptozotocin and 210mg/kg nicotinamide. Then, the diabetes induced rats were treated with N. lappaceum rind at 500mg and 2000mg concentrations for 28 days. Positive control rats were treated with 200mg metformin. In this study, we obtained a 41.06% yield of ethanolic extracts from powdered N. lappaceum rind, while geraniin present in the extract was quantified at 33.0 ± 0.2 mg geraniin/g extract. N. lappaceum rind did not show any toxic effect in the rats both during acute and sub-chronic toxicity study. All biochemical analysis, histology of organs, body weight and relative organ weight clearly showed that there were no significant changes in treated rats. Our study also revealed that, the diabetes induced rats treated with 2000mg N. lappaceum showed reduction in blood glucose level and improved insulin levels which were similar to metformin treated group. Pancreas histology revealed that, the group treated with 2000mg of N. lappaceum had a good distribution of healthy islets and the treatment is comparable to the effect of metformin-treated group. Immunohistochemical staining with Glut-4 and PPAR-gamma antibody also markers for regeneration was over expressed in N. lappaceum treated group with large regenerating islets and the effects were similar to metformin-treated group. In conclusion, N. lappaceum rind extract was able to display anti-hyperglycemic activity at a dose of 2000mg/kg without any toxic effects in high fat diabetes-induced rats.

Rheumatoid arthritis (RA) is a chronic, inflammatory autoimmune disease, where the host immune system attacks the joints. This study used collagen-induced arthritis (CIA) animal model to study the effect of plant-derived antioxidants as the plant-derived antioxidants are known to reduce inflammatory responses in CIA. The aim of the study was to assess the efficacy of ethanol extract from the rind of the rambutan (Nephelium lappaceum) fruit to treat CIA in dark agouti (DA) female rats. Arthritis was induced in DA rats by injecting them with a commercial preparation of collagen from chicken sternal cartilage (4 mg/kg) mixed in complete Freund's adjuvant (CFA). Changes in body weight and joint thickness were monitored every five days from day 0 to 50 of the study. When the signs of arthritis were visible on day 25, the arthritic rats were fed with 300 mg/kg glucosamine or the standardised ethanolic extract (100 or 200 mg/kg) from the rind of the N. lappaceum fruit for 25 days (i.e. until day 50). At autopsy, the sera collected from the animals were tested for C-reactive protein (CRP) and the joints were harvested for histopathology and real-time polymerase chain reaction (qPCR) analysis. iii Both concentrations (100 and 200 mg/kg) of the ethanolic extracts from the rind of the N. lappaceum significantly reduced the arthritis-induced changes in body weight (p<0.05) and paw oedema (p<0.05). There was a significant reduction in the plasma CRP levels (p<0.05) in the treated groups. In addition, a significant reduction (p<0.05) in the arthritis-induced histopathological changes was observed after treatment with the N. lappaceum rind extract. Immunohistochemistry analysis showed that treatment with the N. lappaceum extracts had dose-dependent effects on matrix metalloproteinase-13 (MMP-13) and tissue inhibitor of matrix metalloproteinase-1 (TIMP-1) expression in the joints. There was a minimum of two-fold down regulation (p<0.05) of the TNF-α, COX-2 and NF-κB genes in the joints from arthritic rats treated with the N. lappaceum rind extracts (200 and 100 mg/kg) or glucosamine (300 mg/kg). Treatment with the N. lappaceum rind extracts or glucosamine appear to show regression of RA in the arthritic rats. In conclusion, oral supplementation of standardised ethanolic extract of N. lappaceum rind significantly suppressed pathophysiological, biochemical and histopathological changes induced by collagen administration in the CIA rats. Supplementation with the N. lappaceum extract may be beneficial in preventing the tissue damage and inflammatory conditions in arthritis.

Diminished antioxidant defence or increased production of reactive oxygen species in the biological system can result in oxidative stress which could lead to various neurodegenerative diseases including Alzheimer’s disease (AD). Microglial activation also contributes to the progression of AD by producing several pro-inflammatory cytokines and mediators. Seaweeds have great potential in pharmaceutical and biomedical applications as they are valuable sources of bioactive properties. This study aimed to evaluate the potential of brown (Padina australis and Sargassum polycystum) and green (Caulerpa racemosa) Malaysian seaweeds in inhibiting oxidative stress and neuroinflammation. Anti-oxidative activities of seaweeds were determined by measuring the ability to scavenge 2,2-diphenyl-1-picrylhydrazyl (DPPH) radical, 2,2'-azino-bis(3-ethylbenzothiazoline-6-sulphonic acid) (ABTS) radical and superoxide anion radical. Production of cytokines and PGE2 was measured using enzyme immunoassay and NO was measured using Griess reagent assay. In addition, cytotoxicity assays were conducted using MTT assay. Hexane extract of S. polycystum exhibited potent DPPH radical scavenging ability (IC50 0.16±0.00 mg/mL) and methanol extract of S. polycystum exhibited potent ABTS radical scavenging ability (IC50 0.85±0.02 mg/mL). Hexane extract of C. racemosa showed the strongest superoxide radical inhibitory effect (IC50 0.39±0.01 mg/mL). All extracts of P. australis, S. polycystum and C. racemosa significantly (P<0.05) reduced the production of PGE2, NO, TNF-α, IL-β and IL-6 in a dose dependent manner. At a concentration of α, IL-1β, IL-6) production and approximately more than 50% of NO production. PGE2 secretion was only suppressed by 20-30%. Isolation and structure elucidation of the seaweed compounds were evaluated using mass spectrometer, 1H and 13C Nuclear Magnetic Resonance Spectrometer. Hexane and dichloromethane extracts of S. polcystum and dichloromethane extract of P. australis afforded the same sterol identified as fucosterol and a pigment identified as pheophytin a. Another pigment identified as 132-hydroxy pheophytin a was isolated from methanol extract of S. polycystum. Hexane, dichloromethane and methanol extract of C. racemosa afforded one alkaloid which was identified as caulerpin. Pheophytin a showed the highest DPPH radical scavenging activity with the lowest IC50 value (0.14±0.01mg/mL). All of the seaweed compounds significantly (P<0.05) inhibited the production of PGE2, NO, TNF-α, IL-β and IL-6 in a dose dependent manner. Caulerpin inhibited 60-70% of cytokines, PGE2 and NO. As a conclusion, this study supports the potential application of these seaweeds for the treatment of AD.

The threat of pathogenic infections has almost been eradicated with the introduction of antibiotic treatment regimes. Unfortunately, the repetitive consumption and increasing doses of antibiotics have led to the manifestation of drug tolerance and resistance in microorganisms due to the natural selection process. As a result of this phenomenon, new antibiotics would need to be developed in order to overcome this tolerance/resistance. In this study, antibacterial effects as a result of the preliminary screening between several commercially available essential oils and beta-lactam antibiotics were investigated on multidrug resistant bacteria followed by the modes of action for resistance reversal capability in some of these essential oils. Out of 35 antibiotic-essential oil pairs tested, four showed synergistic effect (FIC ≤ 0.5) and 31 showed no interaction (FIC > 0.5 – 4.0). The preliminary results obtained highlighted the occurrence of a pronounced synergistic relationship between piperacillin/cinnamon bark oil, piperacillin/lavender oil, piperacillin/peppermint oil as well as meropenem/peppermint oil against Escherichia coli J53 R1 and E. coli J53 pMG309 respectively with FIC index in the range 0.26 – 0.5. The time-kill method was used to evaluate the bactericidal activities of essential oils alone or with antibiotics. A rapid and complete killing of bacteria was observed in piperacillin/peppermint, piperacillin/lavender and meropenem/peppermint combinations within 5 – 10 minutes of exposure time. Piperacillin/cinnamon bark combination showed a complete killing profile after 20 h of exposure. Analysis of the membrane permeability effects of essential oils on treated cultures through their stability against sodium dodecyl sulfate demonstrated that essential oils played a role in disrupting the bacterial cell membrane. The action of essential oils on membrane integrity was also confirmed through the decrease in the negative charge of cell surfaces. In addition, the morphology of cells post-treatment with essential oils alone and in combination with antibiotics showed corrugated surfaces and irregular rod-shaped forms under scanning electron microscopic analysis. The overall study emphasised the potential of peppermint, cinnamon bark and lavender essential oils as potential sources of new multi-targeted antimicrobial products. Reduction in the dosage of antibiotics could be employed as a treatment strategy to combat the onset of antibiotic resistance as well as to decrease its adverse effects.

Copper complexes may serve as potential alternatives to platinum-based anticancer agents. By incorporating endogenous metal, such as copper, into metal complexes may reduce toxicity and solve other problems encountered by platinum-based antitumor drugs. A previous study had established that the chiral pairs of ternary copper(II) complex salts, [Cu(phen)(ala)(H2O)]NO3, where ala = alanine, dissociated to yield their respective [Cu(phen)(aa)(H2O)]+ and NO3- ions when dissolved in solution and these species were stable up to more than 24 hours. In this study, two pairs of optically pure chiral [Cu(phen)(ala)(H2O)]X • xH2O, where phen = 1,10-phenanthroline; X = NO3-; ala = L-alanine (L-ala) [LN] and D-alanine (D-ala) [DN]; and X = Cl-; ala = L-ala [LC] and D-ala [DC]; x = number of lattice water molecules, complex salts had been tested on MCF-7 breast cancer cells and its corresponding non-cancer cells, MCF-10A, to investigate the effect of chirality of the alanine and the change in couterion on their anticancer properties and their mechanisms of action. NCI-60 modified MTT assay and morphological study showed that the pairs of chiral ternary copper(II) compounds induced cytostatic and cytotoxic effect in a dose-dependent manner, and there was difference in cell proliferation inhibition on MCF-7 and MCF-10A cells for specific concentration range. The results also showed that chirality did affect their anti-proliferative effect on immortalised MCF-10A but not on cancer cells. However, there were insignificant differences between NO3- and Cl- pairs of the compounds towards MCF-7 and MCF-10A for 48-hour incubation. The ternary copper(II) compounds also induced dose-dependent cell population reduction towards other cancer and non-cancer cell lines, the compounds were selective towards cancer cells over normal cells. Interestingly, human breast carcinoma, MCF-7 was most responsive among the cancer cell lines tested while human hepatocellular carcinoma, HepG2, was the least sensitive towards the ternary copper(II) compounds. The results of screening of [LN] on the NCI panel of 60 human cancer cell lines did show leukaemia was the most resistant and melanoma was the most sensitive. The findings from morphological studies indicated that MCF-7 cells and MCF-10A cells treated with the ternary copper(II) compounds underwent decrease in healthy cells population and cell death by apoptosis. Apoptosis assay using Annexin V-FITC/PI double staining showed that the ternary copper(II) compounds were more selective towards MCF-7 as apoptotic cells in treated MCF-7 cells were more than those in treated immortalised breast MCF-10A cells. Cell cycle data analysis demonstrated that all the compounds suppressed MCF-7 cells growth by accumulate DNA fragments at SubG1 phase. The assay results from using 2’,7’-dicholofluorescein diacetate (DCFH-DA) showed that all the ternary copper(II) compounds induced increase in reactive oxygen species (ROS) in MCF-7 cells with increasing concentration and prolonged exposure (12 to 24 hours) compared to untreated cells. In contrast, the MCF-10A cells, treated under same conditions, showed lower overall ROS generation. The ternary copper(II) compounds induced greater mitochondrial membrane depolarisation in MCF-7 cells than in MCF-10A cells. The effect of all ternary copper(II) compounds on caspase-3/7 activity of MCF-7 cells was no significant for both 12- and 24-hour incubation but they induced distinctive but low increase of caspase-9 activity, especially at 24-hour incubation. In contrast, it was found that treating MCF-10A cells with increasing concentration of each of the ternary copper(II) compounds resulted in activation of both initiator caspase-9 and executor caspase-3/7. These results suggest that apoptosis induction in MCF-10A cells by the above compounds required activation of caspase-3/7 whereas that in MCF-7 did not, thereby implicating activation of different apoptosis pathways. Overall, these findings suggested that ternary copper(II) compounds killed the cancer cells by inducing ROS production, depolarising mitochondrial membrane, and activation of caspase-independent pathway. Therefore, the anticancer properties of the ternary copper(II) compounds involved multiple mode of actions and they also exhibited significant selectivity towards cancer cells rather than non-cancerous cells.

Although many causes have been suggested, the concept of oxidative stress has often been put forward as one of the important causes of neurodegenerative diseases. Natural antioxidants such as flavonoid polyphenols are essential in protecting neuronal cells from undergoing lipid peroxidation and cell death. The present study was designed to elucidate the antioxidant mechanisms of rutin and isoquercitrin in 6-OHDA induced PC12 rat pheochromocytoma cells. Pre-treatment of PC12 cells with rutin or isoquercitrin markedly (p<0.05) reduced the toxic effects caused by exposure to 6-OHDA as this was found to increase the viability of the PC12 cells. In addition, rutin and isoquercetrin were also found to elevate the levels of the endogenous antioxidant enzymes such as superoxide dismutase (SOD), glutathione peroxidase, catalase as well as glutathione. Rutin and isoquercitrin attenuated lipid peroxidation caused by 6-OHDA with p<0.05. Gene expression study using the qPCR array technique showed rutin suppressed the expression of the Parkin genes such Park 5, Park 7 and pro-apoptotic gene Casp3 gene in the PC12 cells exposed to 6-OHDA whilst isoquercitrin pre-treatment down-regulated expression of the Park 5 and Park 7 genes in these cells. Moreover, pre-treatment with rutin stimulated the expression of the TH gene in the PC12 cells. Tyrosine hydroxylase, the protein coded by the TH gene is crucial for biosynthesis of dopamine. Thus, quercetin glycosides treatment may present a novel approach in improving neurodegenerative diseases, including Parkinson’s (PD) and Alzheimer’s (AD) diseases.

The detection of pharmaceuticals in the environment and their impact on the ecosystems have been receiving much interest. Not all the excess pharmaceuticals disposed of by consumers are fully eliminated in the waste water treatment system plants. The final discharge to the environment may contain residual pharmaceuticals which contribute to the burden of chemical pollutants in the receiving waters. The contaminants may have an adverse impact on micro algae, which form the basis of the aquatic food chain. The aim of this study was to assess the toxicity of four pharmaceuticals, namely triclosan, tetracycline, ibuprofen and paracetamol in two microalgae, namely Pseudokirchneriella subcapitata and Dunaliella tertiolecta based on their growth response and cell viability assays, as well as oxidative stress biomarkers such as ROS levels, antioxidant capacity, antioxidant enzymes’ activities and carotenoids. Preliminary growth inhibition tests showed that the sensitivities of both microalgae to the pharmaecuticals were as follows: triclosan > tetracycline > ibuprofen > paracetamol. Triclosan was the most toxic pharmceutical tested, with 96 h EC50 of 42.49 μg/L for P. subcapitata and 10.52 μg/L for D. tertiolecta. The least toxic drug was paracetamol, with 96 h EC50 of 1.96 g/L and 3.33 g/L for P. subcapitata and D. tertiolecta respectively. Triclosan and ibuprofen were chosen for cell viability and oxidative stress biomarker analyses to compare the effect of different classes of drugs in the same microalgae. Cell viability assessed by flowcytometric measurement of cellular fluorescence after staining with fluorescein diacetate (FDA) revealed that the cell viability of P. subcapitata and D. tertiolecta was reduced by approximately 60-90% compared to the control after exposure to ibuprofen, while the cell viability of both microalgae was slightly decreased (5-30%) after exposure to triclosan. The presence of reactive oxygen species (ROS) determined by flowcytometric measurement of cellular fluorescence after staining with 2',7'-dichlorodihydrofluorescein diacetate (H2DCFDA) showed that the percentage of H2DCFDA positive cells increased by 20% when P. subcapitata was exposed to ibuprofen. However, D. tertiolecta did not show any increase in intracellular ROS after being exposed to both toxicants. The total antioxidant capacity of the microalgae determined using DPPH free radical scavenging assay showed that the total antioxidant capacity was significantly increased for D. tertiolecta exposed to triclosan and ibuprofen, while no significant changes were observed for P. subcapitata exposed to both toxicants. Photosynthetic pigments in the microalgae were also affected by pharmaceutical exposure. The carotenoid content in D. tertiolecta increased from 0.4 to 0.5 pg/cell after exposure to ibuprofen, while in P. subcapitata, the carotenoid content decreased from 0.15 to 0.05 after exposure to ibuprofen. Both pharmaceuticals did not cause significant changes in the chlorophyll-a content of D. tertiolecta, while ibuprofen caused severe reduction of chlorophyll-a content in P. subcapitata. Analyses on the antioxidant enzymes’ activities showed that SOD and CAT activities were significantly increased (p < 0.05) in P. subcapitata after exposure to triclosan and ibuprofen. No changes were observed for the SOD enzyme activity in D. tertiolecta exposed to triclosan and ibuprofen, while CAT activity in D. tertiolecta was inhibited by triclosan exposure but enhanced by ibuprofen exposure. In conclusion, the findings suggested that pharmaceuticals could adversely affect the growth and survival of the microalgae tested. Pharmaceuticals could also disrupt the oxidative balance of microalgae and trigger changes in the oxidative stress biomarkers. However, these effects varied between different pharmaceuticals and species of microalgae.

Background: Adequate calcium intake is essential to maximize peak bone mass during the pre-pubertal stage, consecutively supporting bone growth and development as well as preventing bone disorders at Older ages. However, most Malaysian children do not meet calcium requirements from their habitual diets, thus delving into alternative methods for resolving challenges of low calcium intake is crucial. Supplementation with soluble corn fibre (SCF) has been shown to enhance calcium absorption which further improves bone health. The exact mechanism of the action is unclear, which might be due to the production of short-chain fatty acids (SCFAs). As SCFAs are mainly produced from indigestible carbohydrate fermentation by gut microbiome, dietary intake patterns may also play a role in determining SCFA levels and consequently affect bone status. Thus, this secondary data analysis aimed to investigate the association between serum SCFAs level with dietary intake patterns and bone health in Malaysian pre-adolescent children as well as the effect of soluble corn fibre on serum SCFAs level. Methods: A secondary data analysis from the PREBONE-Kids study, with 243 children aged 9-11 were included. Bone mineral density (BMD) and bone mineral content (BMC) were measured using dual-energy X-ray absorptiometry (DXA). Dietary intake was assessed using a 7-days diet history and food intake was further categorised into 23 food groups based on nutrient composition and preparation methods. Dietary intake patterns were then determined based on Malaysian Healthy Eating Index (M-HEI), which consists Of I l components, and Mediterranean Diet Quality Index for Children and Adolescents (KIDMED) questionnaire that comprised Of 16 components. Serum SCFAs were firstly extracted with methanol solvent, followed by measurement of SCFAs using gas chromatography coupled to a (flame-ionization detection detector. Serum SCFA levels were then quantified based on calibration standards. Results: Serum total SCFAs level was inversely correlated with TB BMI) (r (1 1 7) -0.183, p = 0.047). Specific food groups had a significant correlation with serum SCFAs level such as sweetened condensed milk (r (117) = -O. 184, p = 0.046), Sugafr sweetened beverages (r (1 17) = 0.183, p = 0.047), and whole grain (r (1 1 7) = -0.209, p = 0.022). However, dietary intake pattern was not associated with serum SCFAS level (p > 0.05). Significant differences in BMI) and BMC were found among dietary intake pattern groups based on either the M-HEI or KIDMED questionnaire, in which the group with healthier dietary intake patterns had higher bone parameters. Also, KIDMED score had been found to predict BMD (Total body (TB) BMD: = 0.146, p < 0.001; Total body less head (TBLH) BMI): - 0.081,p - 0.008), and BMC (TB BMC: = 0.507, p < 0.001; TBLH BMC: ß = 0.059, p < 0.001) among the study population. There were no significant effect Of SCF on acetic acid (p I .000), propionic acid (p = 1.000), and butyric acid (p = 1.000) over time. Conclusion: The study found that a healthier dietary intake pattern was associated with better BMD and BMC. In addition, although serum SCFAs were associated with specific foods, serum SCFAs were not associated with any specific dietary intake pattern. Also, BMD and BMC were associated with KIDMED score. The supplementation of soluble corn fibre (SCF) did not impact the concentration of serum SCFAs, which amrmed the result Of the PREBONE-Kids study that no added benefit Of SCF supplementation on bone mass. Further research is essential to support the findings as well as to comprehend the mechanism of SCF in the study population. Keywords: Bone health, dietary pattern, pre-adolescent children, short-chain fatty acids, soluble corn fibre

To achieve the objectives, the following was performed and analysed. Bt18 parasporal proteins were separated through anion exchange chromatography and a 68-kDa parasporal protein with cytotoxic activity was purified. Polyclonal IgG (anti-Bt18) for the 68-kDa parasporal protein was successfully raised and purified. The purified anti-Bt-18 IgG showed interaction with parasporal proteins of Bt18, using immunoblot analysis. The purified anti-Bt18 IgG was tested for cross-reactivity against other Bt and Bacillus isolates using Indirect ELISA (quantitative assay). It was observed that Bt isolates share similar antigenic properties with the 68-kDa protein of Bt18 whereas very low cross-reactivity was observed with Bacillus sphaericus, Bacillus subtilis and Bacillus cereus isolates (less than 17%). Immunoblot analysis (qualitative assay) for cross-reactivity also showed that there was a 68- kDa protein in Bt isolates. Cross-reactivity studies via quantitative and qualitative analysis was repeated using purified Bt parasporal protein. Indirect ELISA assay showed the percentage of cross-reactivity was reduced up to 60% after purification for some Bt isolates; Bt2, Bt10, Bt15, Bt19 and Bt20. Meanwhile other Bt isolates still showed more than 90% cross-reactivity. Qualitative assay for cross-reactivity via immunoblot assay detected bands for all the Bt isolates. Similar band profiles were observed in immunoblots using either purified or unpurified Bt parasporal proteins. Receptor binding assay revealed that Bt18 parasporal proteins bound to a 34-kDa protein from the CEM-SS cells lysate. N-terminal amino acid sequence of the 34-kDa protein was GKVKVGVNGFGRIGG. NCBI protein BLAST revealed that the binding protein was Glyceraldehyde-3-phosphate dehydrogenase (GAPDH), a protein with multiple functions, including a vital role in mitochondrial apoptosis. It is interesting to note that Bt18 produces parasporal proteins that act like parasporins. Binding of Bt18 parasporal proteins to GAPDH is a significant finding in this study as literature shows no studies reports on identification of a putative receptor for parasporins. Immuno-staining to detect the binding of Bt18 parasporal protein on CEM-SS cells showed brownish ring formation around the cells, suggesting that the protein may bind to a receptor on the plasma membrane of cells. The specificity of Bt18 parasporal proteins for different monosaccharides showed preferential binding (decreasing order) to D-mannose, D-Lactose and D-glucose. Glycoprotein staining showed that Bt18 parasporal proteins contain three distinct glycoprotein bands about; 200, 100 and 68-kDa. Therefore, two key conclusions can be made; that there is a target protein for Bt18 parasporal proteins in CEM-SS cells and that Bt18 parasporal proteins are lectins that specifically recognise monosaccharides.

Fundamental research on bio-sourced eco-polymers is gaining global interest due to its sustainability, an advantage accepted both in academia and industrial research settings. Malaysia offers rich natural biodiversity, which gives significant opportunities for the application of bio-products and by-products in the medical, biodiesel and pharmaceutical industries. One such bio-product is Ceiba pentandra gum (CPG), a natural product derived from the plant source. CPG exhibits physicochemical characteristics such as conjugation with oppositely charged polyelectrolytes to form polyelectrolytes complexes (PECs). The ability to form PECs provides an opportunity for CPG to be utilised as a potential pharmaceutical excipient in the preparation of controlled-release [CR] dosage forms (Dosage forms that release specified amounts over a stipulated period into the body). Based on our literature search, there has not been any research carried out to investigate the application of CPG as a CR polymer. This research project was aimed at collecting fundamental physicochemical data on CPG, investigate and compare its ability to form PECs with chitosan (CHI), an anionic and polyvinylpyrrolidone (PVP), a neutral polymer. The physicochemical properties of CPG, its PECs formed with CHI and PVP were characterised using a range of fundamental techniques such as pH, conductivity, scanning electromagnetic microscopy, Fourier transform-infrared spectroscopy, powder X-ray diffraction and differential scanning calorimetric (DSC). The ratios of CPG and the polyelectrolytes were optimised to obtain PECs with highest yields using the Design of Experiment (DoE) method employing Response Surface Design concept in the Design Expert 7.0 Database. The optimised PECs were then used to formulate metoprolol succinate CR tablets. The evidence of PECs formation is very conclusive in the DSC study, whereas it was not the case in other studies. The Box Behnken experimental design was used for the optimisation of higher yield of PECs. This resulted in an increased yield of CPG:CHI in the concentration range of 3.11% to 1.04%, within the ratio of 4.86: 1.05. The increased yield was also observed with CPG:PVP in the concentration of 5.03% to 1.12%, within the ratio of 8.45: 1.08. To investigate the drug release retarding efficiency of the CHI and PVP PECs, they were formulated in metoprolol CR tablets. The various tablet formulations met all the in-process standards for tablet preparation. The efficiency of the PECs to retard the drug release was investigated in simulated gastric fluid (SGF) as well as pH 6.8. The release profile of metoprolol at pH 6.8 from the CR tablets containing either CHI or PVP PEC met the dissolution rate standards as stated in United States Pharmacopoeia (USP). However, in the SGF medium, the CHI PEC performed better than PVP PEC in retarding the drug release as per USP dissolution rate standards. In a nutshell, the obtained results provided a platform to use CPG as a CR polymer in the development of once-daily tablet formulation.

Workload-related mental fatigue can have detrimental impact on both work performance and mental health. Saccadic eye movement characteristics (SEMC) have been used to detect mental fatigue however the changes in SEMCs have not been associated to other mental fatigue related measures. This study aims to identify the changes in SEMCs and their associations with mental fatigue-related psychological measures. Participants were required to go through three saccade tasks (pro, anti and mixed) that had elevating difficulty levels. The SEMCs, mental load, and sleepiness were recorded after at each task whilst saliva cortisol levels, fatigue and energy scores were measured pre and post experiment. Data from a total of 55 individuals (aged between 18 to 37 years old) were analysed. Results showed that participants had the highest accuracy rates in the prosaccade task, followed by the mixed task and lastly the antisaccade task. The Visual Analogue Scale-Fatigue (VAS-F) scores showed that there was a slight increase in fatigue and energy in the post-experiment, however, the difference was not statistically significant. However, the participants experienced an increase in mental loading as they progressed from the prosaccade, antisaccade, and to mixed task. From pre and post experiment, the SEMCs were seen to change in the following ways: saccade latency decreased, pupil dilation increased, and peak velocity dropped. In conclusion, the task was able to increase mental loading in participants but may not have been able to significantly induce mental fatigue as suggested by the VAS-F scores. The associations between the psychological measures and SEMCs observed across the three tasks are discussed. The findings of this study can provide a better understanding of the association of saccadic eye movement characteristics with mental fatigue psychological measures, potentially providing insights into new methods for detecting and assessing mental fatigue objectively. Keywords: eye tracking, digital health, saccades, mental load, inhibition, attention

Diabetic foot ulcer (DFU) is one of the most common complications of diabetes mellitus caused by uncontrolled hyperglycemia and typically begins with minor cuts that lead to skin ulceration. Untreated ulcers may develop an infection over time and may persist despite rigorous treatment strategies, and could be due to the presence of biofilms. A biofilm is a layer of bacteria attached strongly to a surface and encased in a tough and slimy substance called extracellular polymeric substance (EPS). This highly virulent bacterial layer contributes to antimicrobial resistance and wound chronicity among DFU patients. This project aims to isolate and identify the bacteria in wounds of DFU patients in Orthopedic Clinic in Serdang Hospital and to investigate their antibiotic sensitivity and their biofilm-forming capabilities. Majority of the patients were Malay of 56-65 years, presenting with Wagner Grade 2 ulcer and had the ulcer for over 4 weeks. Bacteria were isolated from the wound samples and had a high prevalence of Gram-positive bacteria (60.76%). Majority of the bacterial species were able to form biofilms and were most sensitive to imipenem and most resistant to penicillin G. In conclusion, we found that DFU is persistent in elderly male diabetic patients, most of whom only began seeking medical attention for their wound one month from its onset. Bacterial isolation performed on the samples showed that DFU wounds tend to be polymicrobial. The isolates also demonstrate increasing resistance against commonly used antibiotics for treating DFU, which can be worrisome for both clinicians and patients.

Toxoplasma gondii infection is prevalent worldwide and considered one of the neglected parasitic infections targeted by Centers for Disease Control and Prevention. Most healthy individuals suffer no ill effects from toxoplasmosis, despite not undergoing treatment, as the host immunity helps to fight against the infection. However, most of the infected hosts will develop chronic toxoplasmosis as the parasite is capable of encyst itself and hiding themselves from attacks by the host immune system. Hence, understanding of host-parasite metabolic interaction is essential as the parasite strives to survive and grow in the host, while the host struggles to maintain homeostasis in many levels, such as the gut ecosystem, immune system and metabolic system. Here, we have established an animal model using BALB/c mice and Toxoplasma gondii strain ME49, to understand the interaction between both host and parasite using systems biology approaches: metagenomics, metabonomics and host immune response. This study showed a significant and distinctive profile between acute versus chronic phases of infection in terms of host immune system, gut microbiota and urinary metabolic profile. Observation on the change in gut microbe composition occurred during the acute but not chronic phase of infection. Nonetheless, significant changes in host metabolism can still be observed during the chronic phase of infection. Furthermore, our findings also demonstrated perturbation on the host immune responses. Continuous elevation of serum IFN-γ and decrease in levels of TNF-α indicated that IFN-γ plays a vital role in both acute and chronic infection phase of toxoplasmosis. Next, perturbation in the host immune system was also reflected in the urinary metabolic profile. Urinary levels of N-acetyls and O-acetyls of glycoproteins on the termination time point were significantly lower in the infected group. Moreover, infected animal also showed disruption in their host energy metabolism during toxoplasmosis. This was supported by the observation of a decrease in urinary citrate, 2-oxoglutarate and fumarate. Likewise, increase in urinary α-hydroxy N-valerate and β-aminoisobutyrate indicated that disruption of host amino acids catabolism occurred. Lastly, urinary excretion of short-chain fatty acids suggested a transient shift in energy metabolism towards oxidation of both gluconeogenic and ketogenic amino acids such as isoleucine, leucine, valine and nucleotide.

Loss of neurons, excessive accumulation of beta-amyloid (Aβ) peptides in certain brain regions along with accumulation and excessive stimulation of glutamate receptors in the central nervous system (CNS) are common pathological hallmarks of neurodegenerative disease such as Alzheimer’s disease (AD). Although a number of drugs have been approved for the treatment of AD, most of these synthetic drugs have diverse side effects and yield relatively modest benefits. Marine algae have great potential in pharmaceutical and biomedical applications as they are valuable sources of bioactive properties. Hence, this study aimed to provide an overview of potentials of Malaysian seaweeds (Padina australis, Sargassum polycystum, Turbinaria ornata and Caulerpa racemosa) in inhibiting cholinesterase (ChE) enzymes, Aβ-induced and glutamate-induced toxicity. Cholinesterase inhibitory activity was conducted using Ellman’s colorimetric assay while protective effects against Aβ-induced and glutamate-induced toxicity were assessed by determining percentage of viable cells using MTT assay. C. racemosa and S. polycystum showed the most potent anti-acetylcholinesterase activities with the IC50 values ranging from 0.086-0.115 mg/mL. Moreover, C. racemosa and T. ornata were also found to be active against butyrylcholinesterase with IC50 values ranging from 0.118-0.162 mg/mL. Seaweed with dual anti-cholinergic activity is an important finding in this study, as C. racemosa methanol extract and caulerpin exhibited very potent inhibitory activities against both enzymes. Besides that, methanol extract of C. racemosa and caulerpin showed very good neuroprotective effect against beta-amyloid1-42 induced toxicity. In evaluating glutamate-induced toxicity, T. ornata dichloromethane extract had 143.02% viable cells at 0.00030 mg/mL. Moreover, the cell viability continues to increase in the presence of seaweed extracts for other seaweeds extracts except T. ornata extracts. It is extremely noteworthy that the extracts that gave good ChE inhibition activity also gave best protection against Aβ and glutamate induced toxicity. These findings suggest that Malaysian seaweeds have potential to be used as neuroprotective agents in treatment of AD.

Liquid vegetable oils are traditionally ―hardened‖ by partial hydrogenation at high temperature, a process which produces substantial amounts of trans fatty acids (TFAs). TFAs have adverse effects on cardiovascular disease (CVD) risks, food manufacturers have used interesterification as an alternative to harden edible oils. It was reported that stearic rich interesterified (IE) fats may have adverse effects by interrupting the pathway of glucose metabolism, hence elevating plasma glucose and lowering plasma insulin and C-peptide concentration, but yet to be proven in clinical trial. The comparative effects of interesterified fats containing high palmitic acid (C16:0) or stearic acid (C18:0) on CVD risk and insulin resistance have not been well-documented. This study compared the effects of interesterified fats rich in C16:0 obtained from palm olein (IEPOo), with C18:0 from soybean oil (IE Blend) versus that of natural palm olein (POo) on markers of insulin resistance, CVD risks, inflammation effects and obesity in Malaysian adults. In this parallel double-blind long term 8 weeks clinical trial, 50g of test margarine fat incorporated daily into two servings of snacks for breakfast and afternoon tea by three parallel experimental groups, each comprising 28 or 29 adult volunteers aged 20 to 60 years. After 8 weeks, serum triglyceride (TAG) concentration was statistically significantly lower with IE Blend-diet; serum leptin and body fat percentage were also statistically significantly lower in POo-diet compared to IEPOo-diet. No significant changes were found in fasting plasma glucose, serum insulin, C-peptide, serum lipoprotein levels and inflammatory markers across the three test fats. The present study suggest that natural or chemically-interesterified fats formulated margarine rich in either C16:0 or C18:0 do not affect the markers of insulin resistance and inflammation. However, IEPOo fat raised fasting serum TAG, body fat percentage and serum leptin which taken together may contribute to fat deposition.

Alarming numbers of blood stream related nosocomial infections are caused by biofilm formation on catheters by pathogens. This major concern strongly influences prognosis and management of biofilm associated infections, resulting in increased medical costs and death. Antimicrobial coatings over catheters have been effective in reducing microbial adhesion and subsequent biofilm formation. In this study, an attempt was made to coat central venous catheters with antimicrobial-loaded polymeric nanoparticles and evaluate the resulting antibiofilm potential. High Performance Liquid Chromatographic (HPLC) protocols were developed and validated for detection of the two selected antimicrobials targeted for coating the catheters, meropenem and fluconazole. Poly-ɛ-caprolactone polymeric nanoparticles loaded with these antimicrobials were prepared using a modified nanoprecipitation method. With light scattering analyses, meropenem and fluconazole nanoparticles were negatively charged and sized at around 250 nm. Transmission Electron Microscopy (TEM) showed almost spherical nanoparticles. Differential Scanning Calorimetry (DSC) and X-ray diffraction (XRD) analyses indicated drug loading into the polymer and crystalline nature of the nanoparticles. HPLC aided loading studies confirmed loading and identified the best four meropenem and three fluconazole loaded nanoparticles for drug release profiling. Release patterns varied between antimicrobial and among formulations. Inhibitory and cidal concentrations of meropenem against planktonic cells were lower than against biofilm cells and biofilms of Staphylococcus epidermidis and Klebsiella pneumoniae. However, high concentrations of fluconazole were ineffective against planktonic and sessile cells of Candida albicans. A dip-coating technique was then used to coat the catheters by suspending two selected meropenem nanoparticles in a coating solution consisting of ethyl cellulose dissolved in iso-propyl alcohol and triacetin. The coated catheters were then challenged with microbial culture for 72 hours at 37°C with rotary agitation. Biofilms formed over the catheters were quantified with crystal violet staining and microscopically evaluated. Paradoxically, coating with nanoparticles caused roughening of catheter surface and promoted biofilm formation (p<0.05). However, reduction in biofilm formation was observed for Staphylococcus epidermidis (p>0.05) and Klebsiella pneumoniae (p<0.05), when challenged with catheters coated with meropenem nanoparticles. Increasing drug load within the polymeric nanoparticles and further exploration of catheter coating methods are predicted to cumulatively strengthen the antibiofilm ability of antimicrobials-in-nanoparticles coated catheters.

Metal containing polymers have gained much attention in recent years for their antimicrobial properties and few of these polymers are derived from the renewable resources. In view of the growing production of glycerol owing to the implementation of the biodiesel 10% blend (B10) programme in our country, the production of glycerol as the byproduct of the biodiesel is expected to be increased. This study is aimed to synthesise a novel and value-added cobalt-conjugated poly(glycerol adipate) (Co-PGA) with antimicrobial properties for polyurethane coating. Poly(glycerol adipate) (PGA) was first synthesised by polycondensation of glycerol with adipic acid at molar ratio of 2:1 and reaction temperature of 180°C and 210°C to produce PGA with different physico-chemical properties such as OHV, AV, and Mw. Cobalt was then incorporated into PGA at 150 °C and the reactions were monitored by acid value and hydroxyl value determination. The chemical structure and weight average molecular weight of the Co-PGA were characterised by FTIR, 13C-NMR and GPC. The cobalt contents of the synthesised Co-PGA were determined by ICP-MS. PGA retaining with acid value >35 mg KOH/g sample was able to incorporate up to 5%w/w of cobalt into the Co-PGA polymer. The incorporation of cobalt demonstrated additional catalytic effect on producing hyperbranched Co-PGA and increased the degree of branching of the incorporated PGA8 from 25.1 to 59.3%. Co-PGA8 to Co-PGA10 with 4.4-5%w/w of cobalt contents showed inhibition against E. coli (MIC50 at 675 μg/ml), S. aureus (MIC50 at 450 μg/ml ), P. aeruginosa (MIC50 at 112.5 μg/ml), and C. albicans (MIC50 at 56.25μg/ml) in resazurin broth microdilution study. Co-PGA10 incorporated with 5%w/w of cobalt content was used as an active antimicrobial agent to blend with blank PGA and commercial polyols, i.e. PEG 6000 and PCL 2000 and reacted with isophorone diisocyante for the preparation of polyurethane (PU) coating. The antimicrobial properties of the PU coatings prepared by 0.5-35%w/w of the synthesised Co-PGA10 were investigated using disk diffusion test. The PU coatings demonstrated antimicrobial activity against E. coli, S. aureus, B. subtillis and C. albicans. Among all the tested strains, C. albicans was the most susceptible strain to Co-PGA10 and Co-PGA10 prepared PU coating in both broth microdilution and disk diffusion test. The Co-PGA exhibited promising antibacterial and antifungal properties and can be potentially applicable as active antimicrobial agents in protective coating of medical devices.