Antibacterial activity of hexane and methanol fractions of some selected plants against Klebsiella pneumoniae.

Besides adenovirus, pneumonia can also be caused by bacteria. One of the most common bacteria causing the pneumonia is Klebsiella pneumoniae. Currently, treatment by antibiotics has been widely used. Nevertheless, the increasing failure of existing antibiotics because of antibiotic resistance resulted by bacterial pathogens has become a serious problem to human health. Hence, there is a need for a new antibacterial potential agent against K. pneumoniae as an alternative treatment to the pneumonia to prevent the risk of a severe pneumonia for both healthy people and those already infected with the pneumonia. This study, therefore, investigated the antibacterial activity of some selected plants (Pandanus tectorius, Nypa fruticans, Sonneratia alba, Phaleria macrocarpa, Hibiscus tiliaceus, and Pongamia pinnata) against K. pneumoniae. In this study, samples were extracted successively by cold maceration using hexane and methanol. Antibacterial activity was determined by well and disc diffusion methods. Each fraction was prepared by two-fold dilutions from 20 mg/mL to 0.156 mg/mL. All data were analyzed in triplicate replication and presented as mean values ± standard deviation. Results showed that all methanol fractions of selected plants had antibacterial activity against K. pneumoniae, and well-diffusion method showed better antibacterial results compared to the agar well-diffusion method. The strongest activity was obtained by methanol fraction of S. alba leaf, followed by P. pinnata leaf, Nypa fruticans bark, H. tiliaceus leaf, P. macrocarpa leaf, and P. tectorius leaf with the minimum inhibitory concentrations (MICs) value between 0.625 and 5.0 mg/mL. Phytochemical screening revealed that all methanol fractions were rich in flavonoid content, which could have contributed to their antibacterial activity.

The effects of Pandanus tectorius leaf extract on the resistance of White-leg shrimp Penaeus vannamei towards pathogenic Vibrio parahaemolyticus.

Pandanus tectorius leaf extract effect on the White-leg shrimp Penaeus vannamei tolerance against Vibrio parahaemolyticus were investigated in this study. Thirty shrimp post-larvae measured at approximately 1 cm were exposed for 24 h to 0.5, 1, 2, 3, 4, 5 and 6 g/L leaf extract and subsequently observed for survival and immune-related genes expression (Hsp70, ProPO, peroxinectin, penaeidin, crustin and transglutaminase), followed by determination of their tolerance and histological tissue profiles upon Vibrio challenge. Survival of shrimps treated with 6 g/L of leaf extract improved by up to 95% to controls. Hsp70, crustin, and prophenoloxidase mRNA levels were observed to be 8.5, 10.4, and 1.5-fold higher, respectively. Histopathological analysis of the hepatopancreas and the muscle tissues revealed major tissue degeneration in Vibrio-challenged shrimps but not in shrimps primed with P. tectorius leaf extract. Of all the dose examined, the best pathogen resistance results were obtained with a 24 h incubation of shrimp in 6 g/L P. tectorius methanolic leaf extract. The tolerance towards V. parahaemolyticus might be associated with the increased regulation of Hsp70, prophenoloxidase and crustin upon exposure to the extract, all immune-related proteins essential for pathogen elimination in Penaeid shrimp. The present study primarily demonstrated that P. tectorius leaf extract is a viable alternative for enhancing P. vannamei post-larvae resistance against V. parahaemolyticus, a major bacterial pathogen in aquaculture.

Volatile Organic Compounds of Bryophytes from Peninsular Malaysia and Their Roles in Bryophytes.

Volatile emissions from 22 bryophyte species from Peninsular Malaysia were collected using a dynamic headspace technique and analyzed by gas chromatography-mass spectrometry (GC-MS). Thirty organic compounds (VOCs) from eight different groups were detected in bryophytes from the montane forest in Cameron Highlands and the lowland dipterocarp forest in Lata Belatan. The headspace of bryophytes in Cameron Highlands was dominated by tetradecane, 2-ethyl-1-hexanol, decanal, pentanoic acid, 2,2,4-trimethyl-3-carboxyisopropyl, isobutyl ester, D-limonene and naphthalene. On the contrary, in the bryophyte headspace of Lata Belatan, naphthalene and tetradecane were dominant compounds. The elevational pattern detected in VOC composition of bryophytes appears to be an evolutionary feature at the family level that needs verification at other sites. The results also confirmed that the VOC composition of bryophytes is species-specific. The roles of VOCs in bryophytes are presented, including plant-plant communication and plant-insect interaction and as an additional taxonomic character in chemotaxonomy.

Acanthaster planci Inhibits PCSK9 and Lowers Cholesterol Levels in Rats.

Atherosclerosis is the main cause of cardiovascular diseases which in turn, lead to the highest number of mortalities globally. This pathophysiological condition is developed due to a constant elevated level of plasma cholesterols. Statin is currently the widely used treatment in reducing the level of cholesterols, however, it may cause adverse side effects. Therefore, there is an urgent need to search for new alternative treatment. PCSK9 is an enzyme responsible in directing LDL-receptor (LDL-R)/LDL-cholesterols (LDL-C) complex to lysosomal degradation, preventing the receptor from recycling back to the surface of liver cells. Therefore, PCSK9 offers a potential target to search for small molecule inhibitors which inhibit the function of this enzyme. In this study, a marine invertebrate Acanthaster planci, was used to investigate its potential in inhibiting PCSK9 and lowering the levels of cholesterols. Cytotoxicity activity of A. planci on human liver HepG2 cells was carried out using the MTS assay. It was found that methanolic extract and fractions did not exhibit cytotoxicity effect on HepG2 cell line with IC50 values of more than 30 µg/mL. A compound deoxythymidine also did not exert any cytotoxicity activity with IC50 value of more than 4 µg/mL. Transient transfection and luciferase assay were conducted to determine the effects of A. planci on the transcriptional activity of PCSK9 promoter. Methanolic extract and Fraction 2 (EF2) produced the lowest reduction in PCSK9 promoter activity to 70 and 20% of control at 12.5 and 6.25 µg/mL, respectively. In addition, deoxythymidine also decreased PCSK9 promoter activity to the lowest level of 60% control at 3.13 µM. An in vivo study using Sprague Dawley rats demonstrated that 50 and 100 mg/kg of A. planci methanolic extract reduced the total cholesterols and LDL-C levels to almost similar levels of untreated controls. The level of serum glutamate oxalate transaminase (SGOT) and serum glutamate pyruvate transaminase (SGPT) showed that the administration of the extract did not produce any toxicity effect and cause any damage to rat liver. The results strongly indicate that A. planci produced a significant inhibitory activity on PCSK9 gene expression in HepG2 cells which may be responsible for inducing the uptake of cholesterols by liver, thus, reducing the circulating levels of total cholesterols and LDL-C. Interestingly, A. planci also did show any adverse hepato-cytotoxicity and toxic effects on liver. Thus, this study strongly suggests that A. planci has a vast potential to be further developed as a new class of therapeutic agent in lowering the blood cholesterols and reducing the progression of atherosclerosis.

Pandanus tectorius fruit extract promotes Hsp70 accumulation, immune-related genes expression and Vibrio parahaemolyticus tolerance in the white-leg shrimp Penaeus vannamei.

Plants and herbal extracts are indispensable for controlling the spread of disease-causing bacteria, including those that infect aquatic organisms used in aquaculture. The use of plant or herbal extract is expected to be safe for aquatic animals and less harmful to the environment, as opposed to conventional therapeutic alternatives such as antibiotics that promote the occurrence of potential antibiotic-resistant bacteria when used improperly. The efficacy of Pandanus tectorius fruit extract in the regulation of Hsp70 expression, pro-phenoloxidase (ProPO), peroxinectin, penaeidin, crustin and transglutaminase, all immune peptides essential for Vibrio tolerance in white leg shrimp, Penaeus vannamei, was investigated in this study, which included the determination of the safety levels of the extract. Tolerance of shrimp against Vibrio parahaemolyticus, a pathogenic bacteria that causes Acute Hepatopancreas Necrosis Disease (AHPND), was assessed on the basis of median lethal dose challenge survival (LD50 = 106 cells/ml). Mortality was not observed 24 h after exposure of 0.5-6 g/L of the fruit extract, indicating that P. tectorius was not toxic to shrimp at these concentrations. A 24-h incubation of 2-6 g/L of the fruit extract increased shrimp tolerance to V. parahaemolyticus, with survival doubled when the maximum dose tested in this study was used. Concomitant with a rise in survival was the increase in immune-related proteins, with Hsp70, ProPO, peroxinectin, penaeidin, crustin and transglutaminase increased 10, 11, 11, 0.4, 8 and 13-fold respectively. Histological examination of the hepatopancreas and muscle tissues of Vibrio-infected shrimp primed with P. tectorius extract revealed reduced signs of histopathological degeneration, possibly due to the accumulation of Hsp70, a molecular chaperone crucial to cellular protein folding, tissue repair and immune response of living organisms, including Penaeid shrimp.

In vitro and in vivo studies of nanoparticles of chitosan-Pandanus tectorius fruit extract as new alternative treatment for hypercholesterolemia via Scavenger Receptor Class B type 1 pathway.

Pandanus tectorius fruit, a natural product rich in tangeretin and ethyl caffeate, has been reported to have potential as anti-hypercholesterolemia agent via Scavenger Receptor Class B type 1 (SR-B1) pathway. However, due to its semi-polar properties, P. tectorius extract exhibits poor solubility when used as a medical remedy. The extract's solubility can potentially be improved through a synthesis of nanoparticles of chitosan-P. tectorius fruit extract. This can also increase the extract's SR-B1 gene expression activity. To date, no studies of nanoparticles of chitosan-P. tectorius fruit extract and its pathway via SR-B1 have been published anywhere. In this study, cytotoxicity properties against HepG2 were explored by MTT. Then luciferase assay was used to detect their effectiveness in increasing SR-B1 activity. An in vivo study using Sprague dawley was carried out to observe the extract nanoparticles' effectiveness in reducing the cholesterol levels and the toxicity property in rat's liver. As the results showed, the extract nanoparticles had no cytotoxic activity against HepG2 cells and exhibited higher SR-B1 gene expression activity than the non-nanoparticle form. As the in vivo study proved, nanoparticle treatment can reduce the levels of TC (197%), LDL (360%), and TG (109%), as well as increase the level of HDL cholesterol by 150%, in comparison to those for the untreated high-cholesterol diet group. From the toxicity study, it was found that there was non-toxicity in the liver. It can be concluded that nanoparticles of chitosan-P. tectorius fruit extract successfully increased P. tectorius fruit extract's effectiveness in reducing hypercholesterolemia via SR-B1 pathway. Hence, it can be suggested that nanoparticles of chitosan-P. tectorius fruit extract is safe and suitable as an alternative treatment for controlling hypercholesterolemia via SR-B1 pathway.

Inhibitory effects of tangeretin and trans-ethyl caffeate on the HMG-CoA reductase activity: Potential agents for reducing cholesterol levels.

One of the pathways to reduce cholesterol production in the liver is through the inhibition of HMG-Coa reductase (HMGCR) by current drugs, statins. However, these have side effects if consumed in prolonged periods. Tangeretin and trans-ethyl caffeate as alternative drugs in reducing hypercholesterolemia and preventing atherosclerosis have never been reported. Their effects on inhibiting HMGCR activity were investigated through enzymatic method (in vitro and in vivo). The toxicity property was analyzed on the Serum Glutamate Oxalate Transaminase (SGOT)/Serum Glutamate Piruvate Transaminase (SGPT) levels and rat liver histology. The results showed that both compounds inhibited HMGCR activity significantly compare to the control simvastatin (p < 0.05). Tangeretin which showed very good activity in inhibiting HMGCR (83.8 of % inhibition, equal to simvastatin) was selected and used for anti-hypercholesterolemia in vivo assessment. Furthermore, tangeretin was shown to effectively reduced Total Cholesterol (TC) and Low Density Lipoprotein (LDL), and increased High Density Lipoprotein (HDL) levels significantly compared to the simvastatin group (p < 0.05). Tangeretin group was also proven to inhibit HMGCR rat liver activity significantly compare to the control simvastatin (p < 0.05). The toxicity study on the SGOT/SGPT levels and liver histology revealed that there were no side effects after administration by tangeretin. Results found that both tangeretin and trans-ethyl caffeate are potent candidates as anti-hypercholesterolemia agent in vitro. In addition, tangeretin was also shown to be safe and suitable as an alternative treatment for controlling hypercholesterolemia in vivo as well as have potency for preventing atherosclerosis.

Antihypercholesterolemic and antiatherosclerotic potencies of Pandanus tectorius fruits via increasing scavenger receptor-B1 genes expression and inhibition of 3-hydroxy-3-methylglutaryl coenzyme A reductase activity.

Atherosclerosis is a leading cause of death worldwide. The adverse side effects of currently available drugs urge to find more effective and safe remedial agents. Alternative candidates from natural resources are of great consequence in the emerging of new drugs. Pandanus tectorius (Pandanaceae) was traditionally used in Ayurvedic medicine to cure certain diseases. Thus, the current study conducted to elucidate the potency of P. tectorius fruit as antiatherosclerosis and antihypercholesterolemia agents through the regulation of high density lipoprotein (HDL) receptor (scavenger receptor [SR]-B1) gene expression and 3-hydroxy-3-methylglutaryl coenzyme A reductase reductase (HMGCR) in vitro, respectively. The P. tectorius fruit was noncytotoxic against the HepG2 cell line confirmed by 3-(4,5-dimethylthiazol-2-yl) 2,5-diphenyl tetrazolium bromide assay. The P. tectorius fruit successfully upregulates the SR-B1 gene expression and downregulate the HMGCR. Moreover, an in vivo study showed that P. tectorius has good activity on the upregulation of HDL and subsequently downregulation of total cholesterol level. Moreover, P. tectorius fruit did not show any increase in toxicity biomarkers serum glutamic oxaloacetic transaminase and serum glutamate pyruvate transaminase in vivo. These results found that P. tectorius fruits have potency as the preventive agent for hypercholesterolemia and atherosclerosis via SR-B1 and HMGCR mechanisms of action.

Phenolics, fatty acids composition and biological activities of various extracts and fractions of Malaysian Aaptos aaptos

Objective: To investigate phenolics, fatty acids composition and biological activities of various extracts and fractions of Malaysian Aaptos aaptos. Methods: Fatty acid methyl ester was analyzed by gas chromatography-flame ionization detector. Antioxidant activity was determined using 2,2-diphenyl-picrylhydrazyl radical scavenging assay and total phenolics content by Folin-Ciocalteu procedure. Vero cells viability was evaluated using methyl thiazole tetrazolium and the inactivation of herpes simplex virus type 1 by neutral red uptake assay. p-Hydroxybenzamide isolated by column chromatography was characterized by utilizing nuclear magnetic resonance spectroscopy and electron impact mass spectrometry. Results:The chloroform, ethyl acetate and methanol extracts of Aaptos aaptos produced higher portions of straight-chain saturated fatty acid, while hexane extract mainly consisted of unsaturated fatty acid. The five majors of fatty acid methyl ester were identified as behenic acid, cis-10-heptadecenoic acid and cis-10-pentadecenoic acids, palmitic acid and tricosanoic acid. In addition, among all organic extracts, chloroform extract inactivated herpes simplex virus type 1 while exhibited weak cytotoxic activity against normal Vero cells and also exhibited strong cytotoxic activity on HL-60, MCF-7, K562, CEM-SS and WEHI-3B cells. A phenolic compound, p-hydroxybenzamide was also isolated from the sponge. Conclusions: Aaptos aaptos could be a source to derive the potential antiviral and anticancer agents. However, further studies are needed to determine the mechanism involved in the process.

Biological Activities of Isolated Compounds from Three Edible Malaysian Red Seaweeds, Gracilaria changii, G. manilaensis and Gracilaria sp.

Gracilaria species are red marine macroalgae that are found abundantly in Malaysia. Gracilaria changii from Morib, Selangor, G. nanilaensis and Gracilaria sp. from Gelang Patah, Johor were used in this study. Five compounds were successfully isolated and identified as hexadecanoic acid (1), cholest-5-en-3-ol (2), 2-hydroxymyristic acid (3), cholesteryl myristate (4) and 1-(4'-methoxyphenyl)-3-(2",4",6"-trihydroxyphenyl)-3-hydroxypropanone (5) based on spectral data analysis (IR, UV, GC-MS, 'H NMR, "C NMR, HMQC and HMBC). All compounds isolated were tested for cytotoxicity (MTT assay for HL-60 and MCF-7 cell lines), and antibacterial (disc diffusion method), antioxidant (DPPH free radical scavenging assay and xanthine oxidase inhibitory assay) and acetylcholinesterase inhibitory (AChE) activity (TLC bioautographic method). Compounds I and 3 exhibited strong cytotoxic activity against HL-60 and MCF-7 cell lines. Compound 5 showed high antioxidant activity in both the DPPH free radical scavenging and xanthine oxidase inhibition assays. Compound I showed positive activity for AChE inhibitory with a minimum inhibition dose of 0.625 tg sample. All compounds demonstrated antibacterial activity producing 8 to 14 mm inhibition zones. A positive control was applied to all bioassays and experiments were performed with three replicates. Results demonstrated that three edible red seaweeds are rich sources of bioactive compounds with potential application for pharmaceutical purposes.

Phaleria macrocarpa Boerl. (Thymelaeaceae) leaves increase SR-BI expression and reduce cholesterol levels in rats fed a high cholesterol diet.

In vitro and in vivo studies of the activity of Phaleria macrocarpa Boerl (Thymelaeaceae) leaves against the therapeutic target for hypercholesterolemia were done using the HDL receptor (SR-BI) and hypercholesterolemia-induced Sprague Dawley rats. The in vitro study showed that the active fraction (CF6) obtained from the ethyl acetate extract (EMD) and its component 2',6',4-trihydroxy-4'-methoxybenzophenone increased the SR-BI expression by 95% and 60%, respectively. The in vivo study has proven the effect of EMD at 0.5 g/kgbw dosage in reducing the total cholesterol level by 224.9% and increasing the HDL cholesterol level by 157% compared to the cholesterol group. In the toxicity study, serum glutamate oxalate transaminase (SGOT) and serum glutamate pyruvate transaminase (SGPT) activity were observed to be at normal levels. The liver histology also proved no toxicity and abnormalities in any of the treatment groups, so it can be categorized as non-toxic to the rat liver. The findings taken together show that P. macrocarpa leaves are safe and suitable as an alternative control and prevention treatment for hypercholesterolemia in Sprague Dawley rats.