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Our previous study indicated that learning and cognition of obese mice were enhanced by germinated Sang-Yod rice intervention. We recently discover that inferior effects of high fat diet on lipid metabolisms and functions of two vital organs, including the liver and the brain are attenuated by germinated Sang-Yod rice. Thirty-two male C57BL/6J mice are divided into 4 groups (n = 8), and distinctly assigned to receive a different diet for 12 weeks, including normal diet (CONTROL group), high fat diet (HFD group), high fat diet plus 20 mg simvastatin/kg/day (POSITIVE group), and high fat diet plus 0.5% germinated Sang-Yod rice/kg/day (HFD-GR group). Biochemical and histological assessments are performed using blood and tissue samples, respectively. The levels of blood glucose, triglycerides, and high-density lipoproteins for all groups of mice are not different. Increases of low-density lipoprotein and total cholesterol concentrations and the albumin/globulin ratio are found for mice fed with high fat diet. The activities of alanine transaminase and aspartate transaminase enzymes are not different among these groups of mice. Fat degeneration, cytoplasmic vacuoles, and lipid droplets in the livers of mice treated with simvastatin and germinated Sang-Yod rice are reduced. Lastly, cells of the prefrontal cortex of these treated mice are morphologically normal with an absence of microglia. In contrast, microglia is simply observed in this brain area of mice in HFD group. Limitation of high fat in foods should be attended to maintain healthy life.

Health benefits of lady finger were explored in this study by which the ethanol extract from the seeds was utilized for testing. In high glucose (350 mmole/L) medium, the proliferative activity of C2C12 cells was inferior, but their viability was improved following treated with the extract. The activities of antioxidant enzymes, superoxide dismutase (SOD) and catalase (CAT), were also enhanced. In response to such high glucose concentration, glucose uptake ability of these cells was compromised. The extract was demonstrated to increase the ability of glucose uptake by approximately equal to the effect of 200 nmole/L insulin. High intracellular reactive oxygen species (ROS) of RBM-MCS cells grown in high glucose containing medium was elicited. The level of ROS was reduced when the cells were cultured in contact with the extract. L929 cells were made inflammation and oxidative stress by incubating with 37.5 µmole/L DPPH for 30 min. The release of tumor necrosis factor-α (TNF-α) and interleukin 6 (IL-6) from these inflamed cells was inhibited by such treatment. Thus, lady finger might be advantageous in the protection of cells in the body that would be damaged or being susceptible to hyperglycemia and diabetes.

Background A bisresorcinol was isolated as the main constituent of Heliciopsis terminalis’s trunk (Proteaceae). Recently, resorcinol is applied as an active whitening agent in various cosmetic products. Because of the structural mimic to resorcinol, benefits of the bisresorcinol as an aging-enzyme antagonist were demonstrated in this study. Methods The bisresorcinol was purified from the crude ethanolic extract of H. terminalis’s trunk by solvent extraction and preparative chromatography, respectively. Inhibitory activity on collagenase, elastase, and tyrosinase of the compound was investigated by using a different spectroscopic technique. Molecular docking was carried out to predict possible interactions of the substance around the enzyme active sites. Results The IC50 values on collagenase of the bisresorcinol and caffeic acid were 156.7 ± 0.7 and 308.9 ± 1.6 µmole L−1, respectively. For elastase activity, the IC50 of 33.2 ± 0.5 and 34.3 ± 0.3 µmole L−1 was respectively determined for the bisresorcinol and ursolic acid. The bisresorcinol was inhibitory to tyrosinase by exhibiting the IC50 of 22.8 µmole L−1, and that of 78.4 µmole L−1 was present for β-arbutin. The bisresorcinol bound to collagenase, elastase, and tyrosinase with the respective binding energies of −5.89, −5.69, and −6.57 kcal mol−1. These binding energies were in the same ranges of tested inhibitors. The aromatic phenol groups in the structure were responsible for principle as well as supporting binding interactions with enzymes. Hydrogen binding due to hydroxyl groups and π-related attractive forces from an aromatic ring(s) provided binding versatility to bisresorcinol. Conclusion The bisresorcinol purified from H. terminalis might be useful for inclusion in cosmetic products as an aging-enzyme antagonist.

Every part of Calophyllum inophyllum L. has been used in various traditional remedies, especially the oil from its nut was mostly used to treat skin diseases. This study aimed to investigate the composition and antioxidant activity of C. inophyllum nut oil and formulate the oil as a cosmeceutical product. The chemical composition and the amount of total phenolic compounds (TPC) were demonstrated by Gas Chromatograph-Mass Spectrometer (GC-MS) and Folin–Ciocalteu method, respectively. Additionally, the antioxidant activity was tested using the DPPH method. Calophyllolide (4.35%) was a major component. Additional components were calanolide A, inophyllum D, and inophyllum B. We found that the TPC contained 25.9 ± 1.2 mg GE/g oil and a free radical scavenging activity approximate to that of the synthetic Trolox. Emulgel formulation consisted of tween 80, span 80, and isopropyl alcohol as a surfactant, and carbopol 940 as a gelling agent. The microemulsion was formulated using distilled water, oil, tween 80 with span 80, as a surfactant, and isopropyl alcohol as a cosurfactant. The mean droplet size for optimized microemulsion formulations was 34.37 ± 1.06 nm. Furthermore, the results of thermodynamic stability tests (freeze-thaw cycle) and long-term stability tests indicated that emulsions and microemulsions remained stable. In conclusion, this nut oil could potentially be used as a cosmeceutical product, and the obtained emulgels and microemulsions exhibited good characteristics in terms of being a potential agent for skin antioxidant.

Purpose Mesenchymal stem cells (MSCs) are multipotent adult stem cells and present in practically all tissues but originally identified within the bone marrow (BM). The differentiation potential of these cells is generally impaired when culturing in vitro for cell expansion. The aim of this study is to speedily increase the numbers of bone marrow derived mesenchymal stem cells (BM-MSCs) with substantially maintaining their differentiation potential in vitro and improving bone formation in vivo . Materials and methods BM-MSCs isolated from rats were sequentially cultured in α-MEM containing basic fibroblast growth factor (FGF2) and/or insulin to stimulate proliferation and osteogenic commitment, and in the medium with the addition of bone morphogenetic protein 2 (BMP2) and/or bone morphogenetic protein 7 (BMP7) to arouse differentiation. The expression of genes markedly associating the commitment and differentiation were investigated in vitro using real-time PCR technique and mineralization assay, while the capacity of inducing bone formation by the established conditions was determined in vivo using a rat model. Results The BM-MSCs greatly proliferated with active transcription of runx2 and osterix genes when induced by FGF2 and insulin. The in vitro mineralization was enhanced by BMP2, but the extent was diminished when BMP2 was replaced or supplemented by BMP7. Formation of new small blood vessels was notably detected when the cells were respectively challenged by FGF2 plus insulin and BMP2. Conclusion These data are valuable in choosing growth factors for proper bone repair. However, optimization of the established system would be essential when the cells of human source are applied.

Background: Abelmoschus esculentus L. or okra is a vegetable of Malvaceae family. Aim: Okra seeds were dried at 60°C, ground, and extracted by using absolute ethanol. The acquired extract was investigated for chemical constituents, cytotoxicity, anti-tyrosinase activity, and anti-oxidant property. Results: Total phenolic content was equal to 4.6 mg gallic acid/g dry weight, as constituted by tannins, flavonoids, and polyphenols. Saponins and terpenoids were also existent. Based on molecular ion (M+) peaks of GC-MS, long chain fatty acids and their esters (~59%), as well as sterols and their derivatives (~40%) were identified as major components. Glutathione was of 2.5 mg/g dry weight. The anti-tyrosinase activity was equivalent to 15 mg ascorbic acid/g dry weight. The extract was nontoxic to cells of normal fibroblast L929, and exhibited the DPPH radical scavenging activity as well as the ferric reducing antioxidant power, which respectively equivalent to 350 µmol trolox and 170 µmol Fe2+ per g dry weight. Lotions (o/w) containing 1-7% w/w extract were physically and chemically stable over a 6-heating/cooling cycles-period of the stability test. Transdermal penetration of the lotion excipients and active compounds of the extract was not detected according to the Franz-type skin permeation study. Conclusion: Cosmeceuticals containing Abelmoschus esculentus L. seed extract might be useful for whitening effect and to delay skin aging.

Background: Infected bone is often intractable. An ideal approach is to simultaneously eradicate infection and repair the bone defect. The development of osteoinductive bone graft composites to control antibiotic drug release would be useful for the treatment of intractable bone infections. Objectives: To develop a rat model of osteomyelitis for assessing osteoinductive bone graft scaffolds containing antibiotics and a bone morphogenetic protein. Methods: Si-imprinted calcium phosphate is a new hydroxyapatite derivative used in fabricating bone scaffolds. Vancomycin and bone morphogenetic protein-2 (BMP-2) were loaded onto scaffolds of Si-imprinted calcium phosphate using an established method. The efficiency of the scaffold as a drug carrier system was assessed in vivo. Osteomyelitis was induced in rats by infection of the tibial epiphysis with Staphylococcus aureus (BAA 1680). The success of inducing disease was checked after 4 weeks using bacterial culture and radiography. A 10 mm metaphysis bone was surgically removed and replaced with a drug-loaded scaffold. Histology and X-ray imaging were used to evaluate the implants at 8 weeks post implantation. Results: We successfully established a rat model of osteomyelitis. The causative bacteria were effectively eradicated by vancomycin released from the implants. Enhanced bone formation was observed for the implant samples containing vancomycin and BMP-2 compared with those containing either vancomycin or BMP2 alone. Conclusions: The newly developed bone scaffold has potential as a vehicle for therapeutic agents to treat bone diseases.

Silicon-substituted calcium phosphate (Si-CaP) was developed in our laboratory as a biomaterial for delivery in bone tissue engineering. It was fabricated as a 3D-construct of scaffolds using chitosan-trisodium polyphosphate (TPP) cross-linked networks. In this study, heparin was covalently bonded to the residual -NH2 groups of chitosan on the scaffold applying carbodiimide chemistry. Bonded heparin was not leached away from scaffold surfaces upon vigorous washing or extended storage. Recombinant human bone morphogenetic protein 2 (rhBMP-2) was bound to conjugated scaffolds by ionic interactions between the negatively charged SO4 2- clusters of heparin and positively charged amino acids of rhBMP-2. The resulting scaffolds were inspected for bone regenerative capacity by subcutaneous implanting in rats. Histological observation and mineralization assay were performed after 4 weeks of implantation. Results from both in vitro and in vivo experiments suggest the potential of the developed scaffolds for bone tissue engineering applications in the future.

Lactic acid is a product that finds several applications in food, cosmetic, pharmaceutical, and chemical industries. The main objective of this work is to evaluate potential use of the oil palm (Elaeis guineensis) sap as a substrate by Lactobacillus casei. The effects of pH control and nutrient supplement of oil palm sap on fermentation performance were investigated. In this report, when oil palm sap was used as a carbon source for L. casei, pH control did not significantly affect lactic acid production. The addition of de Man, Rogosa, and Sharpe medium improved the biomass and the product yield for which the lactic acid production in static flask at 37 °C and pH 5.5 using 20 g L−1 of total sugar was improved to 0.55 g L−1 h−1. Oil palm sap could serve as a good potential source of raw materials for efficient production of lactic acid by L. casei. When lactic acid was separated from the fermentation broth with Dowex™ 66 resin by water as eluant, the desorption yield was 75.16%. © 2012 Curtin University of Technology and John Wiley & Sons, Ltd.

Background Repairing bone loss by autologous grafting requires that a patient's marrow stromal cells (MSCs) be collected and cultured until the number of cells is adequate for implantation. Currently used techniques allow a slow proliferation rate and produce a culture that contains only small amounts of pluripotent stem cells that will become osteoblasts in culture. Objective To develop culture conditions that permit a rapid increase in the number of MSCs while retaining or improving their potential for complete differentiation in vivo. Results Sequential applications of low doses of basic fibroblast growth factor 2 (FGF2) and bone morphogenetic protein 2 (BMP2) improved the growth and differentiation potential of MSCs. FGF2 also elevated sensitivity of the cells to BMP2. BMP2 increased the syntheses of alkaline phosphatase (ALP), collagen type I and bone sialoprotein, while FGF2 increased the expression of osteocalcin (OC). Full induction as determined by the formation of mineralised nodules in vitro was observed within 7 days. Seeding the induced cells onto scaffolds and then implanting them into nude mice resulted in newly formed bone 4 weeks later. The results of real-time polymerase chain reaction (PCR) and Western blotting suggested that FGF2 increased the pool of committed osteoblasts by up-regulating the Cbfa1/Runx2 gene. The later stages of bone formation seemed to be induced by Cbfa1/Runx2-downstream factors such as BMP2, ALP, collagen type I, bone sialoprotein and OC. Conclusion The culture system that was developed increased both the proliferation of MSC and the proportion that developed into pre-osteoblasts.

Two types of porous ceramic scaffolds were prepared, consisting of β-tricalcium phosphate (TCP) or the mixed powder of TCP and hydroxyapatite (HA) at a 2:1 mass ratio. A variety of methods have been used to fabricate bone scaffolds, while the sintering approach was adopted in this work. An extremely high temperature was used on sintering that proposed to consolidate the ceramic particles. As revealed by SEM, a well opened pore structure was developed within the scaffolds. The θ-values were measured to be of 73.3° and 6.5° for the composite scaffold and TCP sample, respectively. According to XRD patterns, the existence of grains coalescence and partial bonding between HA and TCP powders was demonstrated. Scaffold mechanical property in the term of flexural strength was also determined. The result showed decreasing of the strength by HA supplement, suggesting the more brittle characteristic of HA in comparison with TCP. By soaking the composite scaffold in PBS for a period of 2 weeks, transformation from particles to flank-like crystalline was clearly observed. Such change was found to be favorable for cell attachment, migration, and growth. By implanting cell-seeded scaffolds into nude mice, an abundant osseous extracellular matrix was identified for the composite implants. In contrast, the matrix was minimally detected in TCP implanted samples. Thus, the composite scaffold was found superior for hard tissue regeneration.

New hybrid macroporous scaffolds of polycaprolactone (PCL)/tricalcium phosphate (TCP) were developed by taking into account mechanical properties of the bone to be replaced. FTIR spectra indicated the coating of TCP onto the polymer, providing hydrophilic surfaces necessary for cells to attach. As determined by DSC, the depression of PCL melting point suggested a uniform distribution of PCL within the TCP matrix. SEM micrographs revealed pores of irregular shapes varying from 100-200 µm in size in the resultant structures. Indeed, the pore morphology was precisely determined by the leached particles. The scaffolds could tolerate the impact of at least 5.6 kNm2, making them suitable for use as artificial bones of skull, clavicle and ribs. Rat bone stroma attached and survived on the scaffolds, indicating biocompatible of the used materials. Therefore, the prepared scaffolds would be applicable for bone tissue engineering in the near future.

Aims: To characterize a minimal bacteriocin operon of Prevotella nigrescens ATCC 25261. Methods and Results: A genomic DNA library of Pr. nigrescens ATCC 25261 was constructed and screened for bacteriocin production by an agar overlay assay. Sequence analysis of the bacteriocin-producing recombinant plasmid, pGP2, has shown that the insert DNA consists of 4868 base pairs, termed nig locus. There is a cluster of four genes within the nig locus, respectively designated nigA, B, C and D. Deleting 160 nucleotides at the 3′-end of nigAB resulted in loss of bacteriocin production, indicating that nigAB may belong to a bacteriocin operon. nigA is thought to be the bacteriocin gene, while nigB may encode an immunity protein. Escherichia coli containing pGP2 expressed the bacteriocin, which is similar in size, antimicrobial activity, and biochemical properties to that purified from Pr. nigrescens ATCC 25261. Conclusion: nig Locus is a chromosomal fragment of Pr. nigrescens ATCC 25261, consisting of 4868 base pairs, and has been proved to be important for bacteriocin production. Significance and Impact of the Study: This is the first report of the successful cloning and expression of the bacteriocin from Pr. nigrescens ATCC 25261 into E. coli. This will facilitate the construction of bacteriocin analogues and permit investigation of their structure/function relationships.

Lactic acid is a product that finds several applications in food, cosmetic, pharmaceutical and chemical industries. The main objective of this work is to evaluate potential use of the sap from palmyra ( Borassus flabellifer Linn.) and oil palm ( Elaeis guineensis ) as substrate for lactic acid production by Lactobacillus casei TISTR 1500. The effects of acid hydrolysis, pH control and nutrient supplement of palmyra sap and oil palm sap on fermentation performance were investigated. It was found that lactic acid fermentation using palmyra sap was not significantly affected by either acid hydrolysis or pH control. The addition of MRS increased biomass and product yield. The final lactic acid concentration, dry cell weight and productivity were increased by increasing the total sugars of palmyra sap concentrations up to 134.0 g L -1 . The kinetic parameters for the palmyra sap at 134.0 g L -1 total sugars were calculated to be of: specific growth rate (µ) 0.05 h -1 , the maximum productivity ( R M ) 2.02 g lactic acid L -1 h -1 , cellular yield coefficient ( Y X/S ) 0.20 g cell g -1 sugar, and lactic acid yield ( Y P/S ) 0.78 g g -1 . When oil palm sap was used as carbon source for L. casei TISTR 1500, pH control did not significantly affect lactic acid production. The addition of MRS medium into oil palm sap improved the biomass and the product yield for which the lactic acid production in static flask at 37oC and pH 5.5 using 20 g L -1 of total sugars was improved to be of 0.55 g L -1 h -1 . Oil palm sap could be served as a good potential source of raw materials for efficient production of lactic acid by L. casei TISTR 1500.

To gain insight into how the N-terminal three-stranded beta-sheet-like domain in pediocin-like antimicrobial peptides positions itself on membranes, residues in the well-conserved (Y)YGNGV-motif in the domain were substituted and the effect of the substitutions on antimicrobial activity and binding of peptides to liposomes was determined. Peptide-liposome interactions were detected by measuring tryptophan-fluorescence upon exposing liposomes to peptides in which a tryptophan residue had been introduced in the N-terminal domain. The results revealed that the N-terminal domain associates readily with anionic liposomes, but not with neutral liposomes. The electrostatic interactions between peptides and liposomes facilitated the penetration of some of the peptide residues into the liposomes. Measuring the antimicrobial activity of the mutated peptides revealed that the Tyr2Leu and Tyr3Leu mutations resulted in about a 10-fold reduction in activity, whereas the Tyr2Trp, Tyr2Phe, Tyr3Trp and Tyr3Phe mutations were tolerated fairly well, especially the mutations in position 3. The Val7Ile mutation did not have a marked detrimental effect on the activity. The Gly6Ala mutation was highly detrimental, consistent with Gly6 being in one of the turns in the beta-sheet-like N-terminal domain, whereas the Gly4Ala mutation was tolerated fairly well. All mutations involving Asn5, including the conservative mutations Asn5Gln and Asn5Asp, were very deleterious. Thus, both the polar amide group on the side chain of Asn5 and its exact position in space were crucial for the peptides to be fully active. Taken together, the results are consistent with Val7 positioning itself in the hydrophobic core of target membranes, thus forcing most of the other residues in the N-terminal domain into the membrane interface region: Tyr3 and Asn5 in the lower half with their side chains pointing downward and approaching the hydrophobic core, Tyr2, Gly4 and His8 and 12 in the upper half, Lys1 near the middle of the interface region, and the side chain of Lys11 pointing out toward the membrane surface.

Aims: To characterize the antimicrobial activity produced by Prevotella nigrescens ATCC 25261, and to evaluate its safety on cultured gingival fibroblasts. Methods and Results: An antimicrobial activity was obtained from purifying the culture supernatant of Pr. nigrescens ATCC 25261. Purification of the active compound was achieved with ammonium sulphate precipitation followed by anion-exchange and gel filtration chromatography. As revealed by SDS-PAGE, the active fraction was relatively homogeneous, showing a protein with an approximate molecular weight of 41 kDa. The antimicrobial compound, named nigrescin, exhibited a bactericidal mode of action against Porphyromonas gingivalis, Prevotella intermedia, Tannerella forsythensis, and Actinomyces spp. Nigrescin was stable in a pH range between 6·5 and 9·5, at 100°C for 10 min, and resistant to lyophilization. But its activity was lost after proteinase K treatment. Despite at very high concentrations beyond the minimum inhibitory concentration (MIC), nigrescin was not toxic to the gingival fibroblasts. Conclusion: Nigrescin is a novel bacteriocin produced by Pr. nigrescens ATCC 25261. It exhibits antimicrobial activity against species that are implicated in periodontal diseases. The absence of toxicity on the gingival fibroblasts suggests the possibility in using of nigrescin for an application in periodontal treatment. Significance and Impact of the Study: Novel evidence on nigrescin would make Pr. nigrescens ATCC 25261 attractive in biotechnological applications as an antimicrobial agent in clinical dentistry.

A rapid, specifi c and reliable ion-paired high performance liquid chromatographic assay of small organic acids in bacterial cultures was developed. The assay was conducted using a mixture of 6.0 x 10 -3 mM tetrabutylammonium phosphate and 5 % v/v acetonitrile in water as the mobile phase. Calibration curves for acetic, lactic, and pyruvic acid were linear within the concentration ranges determined (r 2 > 0.9990). Selective removal of interfering compounds from bacterial cultures improved the reliability of the method, demonstrating that more than 90% of the tested acids were recovered. Replicate regression analyses of acid standard plots obtained on three different days gave correlation coeffi cients of r 2 > 0.9995. The assay was precise within a day and between days as indicated by an ANOVA test. According to the validation results and from the analysis of the cultured samples, the described method may be adequate for the routine determination of acid metabolites of lactic acid bacteria.