Targeted Delivery of Lovastatin and Tocotrienol to Fracture Site Promotes Fracture Healing in Osteoporosis Model: Micro-Computed Tomography and Biomechanical Evaluation

Osteoporosis is becoming a major health problem that is associated with increased fracture risk. Previous studies have shown that osteoporosis could delay fracture healing. Although there are potential agents available to promote fracture healing of osteoporotic bone such as statins and tocotrienol, studies on direct delivery of these agents to the fracture site are limited. This study was designed to investigate the effects of two potential agents, lovastatin and tocotrienol using targeted drug delivery system on fracture healing of postmenopausal osteoporosis rats. The fracture healing was evaluated using micro CT and biomechanical parameters. Forty-eight Sprague-Dawley female rats were divided into 6 groups. The first group was sham-operated (SO), while the others were ovariectomized (OVx). After two months, the right tibiae of all rats were fractured at metaphysis region using pulsed ultrasound and were fixed with plates and screws. The SO and OVxC groups were given two single injections of lovastatin and tocotrienol carriers. The estrogen group (OVx+EST) was given daily oral gavages of Premarin (64.5 µg/kg). The Lovastatin treatment group (OVx+Lov) was given a single injection of 750 µg/kg lovastatin particles. The tocotrienol group (OVx+TT) was given a single injection of 60 mg/kg tocotrienol particles. The combination treatment group (OVx+Lov+TT) was given two single injections of 750 µg/kg lovastatin particles and 60 mg/kg tocotrienol particles. After 4 weeks of treatment, the fractured tibiae were dissected out for micro-CT and biomechanical assessments. The combined treatment group (OVx+Lov+TT) showed significantly higher callus volume and callus strength than the OVxC group (p<0.05). Both the OVx+Lov and OVx+TT groups showed significantly higher callus strength than the OVxC group (p<0.05), but not for callus volume. In conclusion, combined lovastatin and tocotrienol may promote better fracture healing of osteoporotic bone.     

Evidence of geographical structuring in the Malaysian Snakehead, Channa striata based on partial segment of the CO1 gene

Channa striata, locally known as "haruan", is economically important in fisheries and aquaculture industries in several Asian countries. DNA sequencing, based on a partial segment of the Cytochrome oxidase c subunit 1 (CO1) gene, was used to determine genetic variation in C. striata samples from four different populations on the west coast of Peninsular Malaysia. The highest nucleotide and haplotype diversities were observed in the Linggi population (π = 0.0067, h = 0.835), and the lowest in the Timah Tasoh population (π = 0.0008, h = 0.286). Apart from Kajang-Linggi, which was insignificant, F(ST) values were significant (p < 0.05) in all pairwise-population comparisons. Consequently, it is inferred that genetic structuring C. striata populations in this region was largely shaped by a common origin, with secondary influences from geographical factors and isolation.     

Genetic structure of the snakehead murrel, Channa striata (channidae) based on the cytochrome c oxidase subunit I gene: Influence of historical and geomorphological factors

Nucleotide sequences of a partial cytochrome c oxidase subunit I gene were used to assess the manner in which historical processes and geomorphological effects may have influenced genetic structuring and phylogeographic patterns in Channa striata. Assaying was based on individuals from twelve populations in four river systems, which were separated into two regions, the eastern and western, of the biodiversely rich state of Perak in central Peninsular Malaysia. In 238 specimens, a total of 368-bp sequences with ten polymorphic sites and eleven unique haplotypes were detected. Data on all the twelve populations revealed incomplete divergence due to past historical coalescence and the short period of separation. Nevertheless, SAMOVA and F(ST) revealed geographical structuring existed to a certain extent in both regions. For the eastern region, the data also showed that the upstream populations were genetically significantly different compared to the mid- and downstream ones. It is inferred that physical barriers and historical processes played a dominant role in structuring the genetic dispersal of the species. A further inference is that the Grik, Tanjung Rambutan and Sungkai are potential candidates for conservation and aquaculture programmes since they contained most of the total diversity in this area.     

Andrographis paniculata ameliorates carbon tetrachloride (CCl(4))-dependent hepatic damage and toxicity: diminution of oxidative stress

Andrographis paniculata (hempedu bumi) is a plant that possesses many medicinal values in treating several diseases and for health care maintenance. However, its hepatoprotective activity and mechanism of action have not been fully investigated. Therefore, this study aimed to evaluate the hepatoprotective effects of A. paniculata and its mechanism of action in rats. Carbon tetrachloride (CCl(4)) challenge of rats at a dose of 1.2 ml/kg body weight-induced oxidative stress in the liver. This was evidenced by augmentation in lipid peroxidation, which was accompanied by a decrease in the activities of antioxidant enzymes and depletion in the level of reduced glutathione (P < 0.05). Parrallel to these changes, CCl(4) challenge too, enhanced hepatic damage as evidenced by sharp increase in serum transaminases (e.g. alanine aminotransferase, aspartate aminotransferase, and lactate dehydrogenase) (P < 0.05). Additionally, the impairment of liver function corresponded to histolopathological changes. However, most of these changes were reversed in a dose-dependent fashion by pre-treatment of animals with A. paniculata (P < 0.05). The ability of A. paniculata to scavenge the 2,2-Diphenyl-2-picrylhydrazyl radical was determined through its EC(50) value. The EC(50) value of A. paniculata was 583.60 ± 4.25 μg/ml. In addition, A. paniculata was found to contain 65.37 ± 1.20 mg/g total phenolics expressed as gallic acid equivalent. From these studies, it is concluded that A. paniculata could be used as a hepatoprotective agent and possesses the potential to treat or prevent degenerative diseases where oxidative stress is implicated.     

Herbivore- and Elicitor- Induced Resistance in Groundnut to Asian armyworm,Spodoptera litura (Fab.) (Lepidoptera: Noctuidae)

Induced defense was studied in three groundnut genotypes ICGV 86699 (resistant), NCAc 343 (resistant) and TMV 2 (susceptible) in response to Spodoptera litura infestation and jasmonic acid (JA) application. The activity of the oxidative enzymes [peroxidase (POD) and polyphenol oxidase (PPO)] and the amounts other host plant defense components [total phenols, hydrogen peroxide (H2O2), malondialdehyde (MDA), and protein content] were recorded at 24, 48, 72 and 96 h in JA pretreated (one day before) plants and infested with S. litura, and JA application and simultaneous infestation with S. litura to understand the defense response of groundnut genotypes against S. litura damage. Data on plant damage, larval survival and larval weights were also recorded. There was a rapid increase in the activities of POD and PPO and in the quantities of total phenols, H2O2, MDA and protein content in the JA pretreated + S. litura infested plants. All the three genotypes showed quick response to JA application and S. litura infestation by increasing the defensive compounds. Among all the genotypes, higher induction was recorded in ICGV 86699 in most of the parameters. Reduced plant damage, low larval survival and larval weights were observed in JA pretreated plants. It suggests that pretreatment with elicitors, such as JA could provide more opportunity for plant defense against herbivores.     

Role of salicylic acid in induction of plant defense system in chickpea (Cicer arietinum L.)

Salicylic acid (SA), a plant hormone plays an important role in induction of plant defense against a variety of biotic and abiotic stresses through morphological, physiological and biochemical mechanisms. A series of experiments were carried out to evaluate the biochemical response of the chickpea (Cicer arietinum L.) plants to a range of SA concentrations (1, 1.5, and 2 mM). Water treated plants were maintained as control. Activities of peroxidase (POD) and polyphenol oxidase (PPO) were evaluated and amounts of total phenols, hydrogen peroxide (H₂O₂), and proteins were calculated after 96 h of treatment. Plants responded very quickly to SA at 1.5 mM and showed higher induction of POD and PPO activities, besides the higher accumulation of phenols, H₂O₂ and proteins. Plants treated with SA at 2 mM showed phytotoxic symptoms. These results suggest that SA at 1.5 mM is safe to these plants and could be utilized for the induction of plant defense.     

LC-MS/MS-based metabolites of Eurycoma longifolia (Tongkat Ali) in Malaysia (Perak and Pahang)

A number of three LC-MS/MS hybrid systems (QTof, TripleTof and QTrap) has been used to profile small metabolites (m/z 100-1000) and to detect the targeted metabolites such as quassinoids, alkaloids, triterpene and biphenylneolignans from the aqueous extracts of Eurycoma longifolia. The metabolite profiles of small molecules showed four significant clusters in the principle component analysis for the aqueous extracts of E. longifolia, which had been collected from different geographical terrains (Perak and Pahang) and processed at different extraction temperatures (35°C and 100°C). A small peptide of leucine (m/z 679) and a new hydroxyl methyl β-carboline propionic acid have been identified to differentiate E. longifolia extracts that prepared at 35°C and 100°C, respectively. From the targeted metabolites identification, it was found that 3,4?-dihydroeurycomanone (quassinoids) and eurylene (squalene-type triterpene) could only be detected in the Pahang extract, whereas canthin-6-one-3N-oxide could only be detected in the Perak extract. Overall, quassinoids were present in the highest concentration, particularly eurycomanone and its derivatives compared to the other groups of metabolites. However, the concentration of canthin-6-one and β-carboline alkaloids was significantly increased when the roots of the plant samples were extracted at 100°C.     

Three phases hollow fiber LPME combined with HPLC-UV for extraction, preconcentration and determination of valerenic acid in Valeriana officinalis

In the present work, the applicability of hollow fiber-based liquid phase microextraction (HF-LPME) was evaluated for the extraction and preconcentration of valerenic acid prior to its determination by reversed-phase HPLC/UV. The target drug was extracted from 5.0 mL of aqueous solution with pH 3.5 into an organic extracting solvent (dihexyl ether) impregnated in the pores of a hollow fiber and finally back extracted into 10 μ L of aqueous solution with pH 9.5 located inside the lumen of the hollow fiber. In order to obtain high extraction efficiency, the parameters affecting the HF-LPME, including pH of the donor and acceptor phases, type of organic phase, ionic strength, the volume ratio of donor to acceptor phase, stirring rate and extraction time were studied and optimized. Under the optimized conditions, enrichment factor up to 446 was achieved and the relative standard deviation (RSD) of the method was 4.36% (n = 9). The linear range was 7.5-850 μg L?1 with correlation coefficient (r2=0.999), detection limits was 2.5 μg L?1 and the LOQ was 7.5 μg L?1. The proposed method was evaluated by extraction and determination of valerenic acid in some Iranian wild species of Valerianaceae.     

Improved mannan-degrading enzymes' production by Aspergillus niger through medium optimization

The effect of different carbon and nitrogen sources on the production of mannan-degrading enzymes, focussing on β-mannanase, by Aspergillus niger was investigated using shake flask culture. The β-mannanase activity obtained during growth of A. niger on guar gum (GG, 1495 nkat mL(-1)) was much higher than those observed on other carbon substrates, locust bean gum (1148 nkat mL(-1)), α-cellulose (10.7 nkat mL(-1)), glucose (8.8 nkat mL(-1)) and carboxymethylcellulose (4.6 nkat mL(-1)). For fermentation using GG as a carbon source, bacteriological peptone gave the highest β-mannanase activity (1744 nkat mL(-1)) followed by peptone from meat (1168 nkat mL(-1)), yeast extract (817 nkat mL(-1)), ammonium sulphate (241 nkat mL(-1)), ammonium nitrate (113 nkat mL(-1)) and ammonium chloride (99 nkat mL(-1)) when used as a nitrogen source. The composition of bacteriological peptone and initial pH of the medium were further optimized using response surface methodology (RSM). Medium consisted of 21.3 g L(-1) GG and 57 g L(-1) peptone with initial culture pH of 5.5 was optimum for β-mannanase production (2063 nkat mL(-1)) by A. niger. The β-mannanase production obtained in this study using A. niger was significantly higher than those reported in the literature.