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DOI: 10.14457/cu.the.2007.1599
title: Gene expression analysis of russell's viper venom and molecular cloning, expression and functional study of RV factor X activator

description: Russell's viper, Daboia russelli, is a venomous snake widely distributed in several parts of Asia. RV bites from different geographical areas cause variety of signs and symptoms with severe injury or even death in human. This study was aim at (1) characterization the toxin composition of Thai RV venom by Expressed Sequence Tags (ESTs) analysis, (2) utilization of the medical useful toxin by DNA recombinant technology, and (3) production of toxin-specific antibody by DNA immunization. ESTs are an important and rapid tool to characterize the pattern of gene expression of organism. cDNA library was constructed from the snake venom glands to study the active genes in the venom. A total 135 ESTs have been generated from size-selected clones at random. Fifty-seven percents (77/135) of ESTs were significant matching to the entries in the database. Of these, several ESTs involved in the toxicity of the venom were obtained, e.g. phospholipase A2, factor X activator and factor V activator, BPP-CNP homolog, etc. Some of cDNA sequences of these genes showed a variation from the other reports from RV from other region. These ESTs data could provide a resource of the expressed pattern relating to the clinical pathology of RV bite. In the mean time, factor X activator (RVV-X), one of RV key toxin, was cloned, characterized, and expressed in Pichia pastoris. Northern analysis indicated that the mRNA of RVV-X heavy chain and light chain were about 2.5 kb and 1.0 kb, respectively. However, the expression of RVV-X was unappreciated in Pichia yeast. Only heterologous RVV-X light chain could be seen in SDS-PAGE while RVV-X heavy chain could not be detected. RVV-X light chain could also be detected by anti-His (C-term) antibody and RVV-immunized horse antibodies. Moreover, to generate toxin-specific antibody, RVV-X was cloned into pVaxSec mammalian expression vector for DNA immunization in mice. RVV-X disintegrin domain showed to be a good DNA immunogen in antibody production in mice while RVV-X with other domains gave unsatisfied results. Antibody from RVV-X disintegrin domain-immunized mouse serum could bind to the RVV-X in both reduced and non-reduced forms. In summary, this work has identified the Thai RV venom composition by characterization the RV transcripts and could produce heterologous RVV-X light chain in Pichia pastoris. In addition, RVV-X-specific antibody was successfully produced by DNA immunization in mice which may be developed for the large-scale antibody production in other mammals
DOI: 10.14457/cu.the.2007.1600
title: Transesterification of palm oil using base loaded ERB-1 catalysts for biodiesel production

description: ERB-1 was hydrothermally synthesized by rotating crystallization and using piperidine (PD) as a structure directing agent with the gel composition 1.50 SiO2 : B2O3 : 0.60 NaOH : 1.80 PD : 28.50 H2O. The gel was crystallized at 175oC for 7 days and removed template from as-synthesized sample by calcination in a muffle furnace at 550°C. The del-ERB-1 was prepared by swelling and exfoliating the ERB-1 precursor. The del-ERB-1 consists of thin sheet with an extremely high external surface area. NaERB-1 and Na-del-ERB-1 catalysts could be obtained by stirring with 0.10 M NaOH solution at RT for 3 h and this condition was applied to load with other alkali ions (K, Rb and Cs). All catalysts were characterized using X-ray diffraction, inductively coupled plasma atomic emission, nitrogen adsorption and scanning electron microscopy and tested catalytic activity in transesterification reaction of palm oil with methanol to produce fatty acid methyl esters (biodiesel). The various reaction conditions such as catalyst amount, methanol to oil mole ratio including reaction time and temperature were studied. The product was analyzed by GC technique. In case of del-ERB-1 catalyst, it shows the triglyceride conversion slightly higher than non-delaminated one (2.28%), while conversion over Na-del-ERB-1 is lower than NaERB-1 (4.49%). Thus, the highest transesterification activity is achieved when NaERB-1(0.10, RT) was used as catalyst, the conversion and methyl esters yield can be reached to 74.03% and 42.45%, respectively. Moreover, catalytic activities of used and Na-reloaded catalysts were also investigated. The activity of Na-reloaded ERB-1 catalyst is slightly higher than used ERB-1 but lower than the fresh one.
DOI: 10.14457/cu.the.2007.1601
title: Devise of a Cleaning in Place (CIP) with a circulation process in a pasteurized milk tank

description: Cleaning In Place (CIP), one of cleaning processes which is widely used in most food and beverage industrials, has been applied to clean raw materials and products remained in storage tanks, pipe lines and equipment because the CIP process provides reliable, fast and efficient cleaning without dismantling equipment. Although, the CIP cleaning process have been developed continuously to increase chemical and water saving, the real operations of these processes still have been considered as a large amount consumed water processes. This work studies process behaviors and optimizes to determine optimal draining ratio of the CIP cleaning chemicals in a dairy plant. To achieve these, mathematical models of the CIP process have been developed and data have been gathered from real plant. With these models, simulation study has been carried out to find out the dynamic behavior of the process with respect to the concentrations and contaminations in cleaning chemicals. The optimization problem has been formulated and solved using written programs based on MATLAB application program. According to the microbial data result, the used cleaning detergent, alkaline and acid, can be recycled. In the optimization approach, the optimal draining ratios have been obtained as 4.34% and 1.70% with 2 and 3 cycles/time draining time after 1 week pass for alkaline and acid respectively.
DOI: 10.14457/cu.the.2007.1602
title: Chemical constituents and their biological activities of Gymnema inodorum Decne. and Moringa oleifera Lam

description: The CH [subscript 2]Cl[subscript 2] and n-BuOH crude extracts of G. inodorum and those of the roots of M. oleifera revealed a promising antioxidant and α-amylase inhibitor activities. Chromatographic separation of the CH[subscript 2]Cl[subscript 2] extract of G. inodorum led to the isolation of three known substances: a long chain alcohol, a long chain ketone and a mixture of steroids containing stigmasterol as a major component. While two known saponins, (3β,4α,16β)-16,23,28-trihydroxyolean-12-en-3-yl-β-D-glucopyranosiduroic acid and (3β,16β,22α)-16,28-dihydroxyolean-12-en-3-yl-O-β-D-glucopyranosyl-β-D-glucopyranosiduroic acid were isolated from the n-BuOH crude extract. Four known substances: a long chain alcohol, a long chain ketone, a mixture of β-sitosterol, campesterol and stigmasterol along with a mixture of triglycerides as 1,3-dilinoleoyl-2-olein, 1,3-dioleoyl-2-linolein and 1,2,3-trilinolein were isolated from the CH [subscript 2]Cl[subscript 2] crude extract of the roots of M. oleifera. In addition, five known compounds were isolated and identified as linoleic acid, isothiocyanatomethylbenzene, 1,2,3-triolein, benzamide, β-sitosteryl linoleate and β-sitosterol-3-O-β-D-glucopyranoside. All isolated substances were assayed for antioxidant, the inhibition of cholinesterase activity and α-amylase inhibitor. Only (3β,16β,22α)-16,28-dihydroxyolean-12-en-3-yl-O-β-D-glucopyranosyl-β-D-glucopyranosiduroic acid was active for radical scavenging effect on DPPH radical. The α-amylase inhibition of (3β,16β,22α)-16,28-dihydroxyolean-12-en-3-yl-O-β-D-glucopyranosyl-β-D-glucopyranosiduroic acid (38.59%) was better than that of (3β,4α,16β)-16,23,28-trihydroxyolean-12-en-3-yl-β-D-glucopyranosiduroic acid (9.80%). Moreover, benzamide isolated from the CH2Cl2 crude extract of M. oleifera roots showed the highest scavenging activity on DPPH, while β-sitosterol-3-O-β-D-glucopyranoside was found as a good inhibitor for α-amylase inhibition with IC50 of 0.326 mM. The synthetic derivatives of certain carboxamides and esters were also prepared to test for scavenging effects on DPPH radical. N-(2-hydroxylphenyl)-5-(3,4-methylenedioxyphenyl)-2E,4E-pentadieneyl amide showed highly active toward radical scavenging effect on DPPH radical with IC50 of 0.470 mM and ABTS value being similar to that of trolox revealing good potent for antioxidant activity.
DOI: 10.14457/cu.the.2007.1603
title: Limbal stem cells maintenance in the in vitro cultivation

description: The maintenance of a healthy corneal epithelium under both normal and wound healing conditions is achieved by a population of stem cells (SC) located in the basal epithelium at the corneoscleral limbus. These cells, also called limbal epithelial stem cells (limbal SC) retain their self-renewal and maintain their proliferative capacity. Limbal stem cell can be expanded ex vivo on amniotic membrane and used for transplantation to the patients who suffer from limbal stem cell deficiency. However, this technique still has drawbacks; particularly stem cells lose their properties that limit the use for transplantation. Therefore, the studying about limbal stem cells niche and developing a culture system that similar to their niche are required for maintenance limbal stem cell in long term culture by activating Wnt signaling in limbal stem cells and creating Bone morphogenesis protein (BMP) antagonists expression feeder cells. These molecules have been showed a potential role in the proliferation and the maintenance of other epithelial stem cells in their niche. We founded BMP antagonist, Gremlin-1, prolonged cell cycle of limbal stem cell through down-regulation of cyclin-dependent kinase inhibitor, p27 [subscript kip1] and activation of Wnt signaling promoted proliferation of limbal stem cells.
DOI: 10.14457/cu.the.2007.1604
title: Neotectonics along the Pua fault in Changwat Nan, Northern Thailand : evidence from remote sensing and thermoluminescence dating
DOI: 10.14457/cu.the.2007.1605
title: The impacts of political events on stock return and volatility : case of Thailand

description: This study investigates the stock return and volatility impact of various Thai political events on different group of firms based on four characteristics, which are foreign ownership structure, growth options, analyst coverage and financial status. The sample period covers 1996 to 2006 using daily data of all Thai listed firms. The results suggest that political news does not affect all firms identically, also the impact is better observed when political news are separately studied by their type. Political news related to government changes of authority is correlated to positive returns, while, domestic- conflict related news is correlated to large negative returns. At firm level, the study finds positive response from high growth stocks and high analyst recommended stocks towards news representing less degree of uncertainty, i.e., the government changes of authority. However, their negative response on domestic-conflict events was even worse than low growth stocks and low analyst recommended stocks. Financially weak firms are the least favorable among all groups of firms on the event date with domestic conflicts. Volatility impact is observed in some portfolios for specific type of news. The study, therefore, gives insight into the forces of Thai political events that varies from firm to firm.
DOI: 10.14457/cu.the.2007.1606
title: Characteristics and catalytic properties of boron-modifed zirconia-supported cobalt catalyst for carbon monoxide hydrogenation

description: In the present study, differences in characteristics and catalytic activity towards CO hydrogenation of Co catalysts dispersed on the various B modification on zirconia supports were investigated. After calcination, the various samples were characterized using XRD, SEM/EDX, TEM, TPR and H2 chemisorption. CO hydrogenation (H₂/CO = 10/1) was also performed to determine the overall activity and selectivity. The B modification on zirconia supports was found to increase the catalytic activities of Co catalyst (20 wt%) during CO hydrogenation. The amounts of B loading in the zirconia support were in the range of 0.5 to 3 wt%. The increased activities were attributed to the larger number of reduced Co metal surface atoms for catalyzing the reaction. It is worth noting that B modification can result in (i) preventing the agglomeration of Co oxide species, and (ii) increasing the dispersion of Co oxide species leading to facilitating the reduction of Co oxide species into Co metal surface atoms. In addition, only a slight decrease in C₂-C₄ selectivity was observed. For the various cobalt loading ( 5 to 20 wt%) on the 1 wt % of boron modification on zirconia, it was found to decrease the catalytic activity with decreasing the amount of cobalt loading, but an increase in C₂-C₄ selectivity was observed.
DOI: 10.14457/cu.the.2007.1607
title: Antiviral activity against herpes simplex virus of lipoic acid

description: การศึกษาฤทธิ์ต้านไวรัสเฮอร์ปีส์ซิมเพล็กซ์ทัยป์ 1 สายพันธุ์ KOS และทัยป์ 2 สายพันธุ์ Baylor 186 ของ lipoic acid และ lipoamide ใน Vero cells โดยวิธี inactivation, plaque reduction และ MTT reduction assay พบว่า lipoic acid และ lipoamide มีฤทธิ์ต้าน HSV-1 และ HSV-2 โดยค่าความเข้มข้นของสารที่ยับยั้งการเจริญของไวรัส 50% (IC50 ) ของ lipoic acid ในการยับยั้ง HSV-1 เท่ากับ 96.34, 115.49 และ 126.85 ug/ml ตามลำดับ และมีค่า SI เท่ากับ 3.04, 2.54 และ 2.31 ค่า IC50 ของ lipoic acid ในการยับยั้ง HSV-2 เท่ากับ 111.75, 130.13 และ 135.06 ug/ml ตามลำดับ โดยมีค่า SI เท่ากับ 2.62, 2.25 และ 2.17 ค่า IC50 ของ lipoamide เท่ากับ 41.28, 60.47 และ 105.17 ug/ml สำหรับ HSV-1 และ 50.15, 83.49 และ 108.42 ug/ml สำหรับ HSV-2 ตามลำดับ โดยมีค่า SI ในการยับยั้ง HSV-1 เท่ากับ 6.57, 4.48 และ 2.58 ซึ่งสอดคล้องกับค่า SI ในการยับยั้ง HSV-2 ที่มีค่าเท่ากับ 5.41, 3.25 และ 2.50 การทดสอบความเป็นพิษต่อเซลล์ของ lipoic acid และ lipoamide โดยวิธี trypan blue exclusion และ MTT reduction assay พบว่าความเข้มข้นของสารที่เป็นพิษต่อเซลล์เพาะเลี้ยง 50% (CC50 ) มีค่าเท่ากับ 242.69 และ 292.19 ug/ml สำหรับ lipoic acid และ 239.03 และ 271.10 ug/ml สำหรับ lipoamide ตามลำดับ นอกจากนี้ในการประเมินกลไกการออกฤทธิ์เบื้องต้นของ lipoic acid ในการยับยั้ง HSV โดยใช้วิธี virucida, post-binding, penetration, prophylactic activity และ virus growth inhibition assay พบว่า lipoic acid ไม่มีฤทธิ์ยับยั้งการจับของ HSV บนผิวเซลล์เพาะเลี้ยง ไม่มีฤทธิ์ป้องกันการเข้าสู่เซลล์ของไวรัสและไม่มีฤทธิ์ทำลายอนุภาคของไวรัสโดยตรง อย่างไรก็ตาม lipoic acid มีผลลดปริมาณ HSV-1 และ HSV-2 ใน Vero cells ได้มากกว่า 70% โดยเฉพาะอย่างยิ่งใน lopoic acid มีผลลดปริมาณ HSV-1 และ HSV-2 ใน Vero cells ได้มากกว่า 70% โดยเฉพาะอย่างยิ่งใน pre-treatment นอกจากนี้ lipoic acid ยังแสดงฤทธิ์ต้าน HSV ในเซลล์เพาะเลี้ยงชนิดอื่น เช่น human cervix epithelium HeLa cells และ normal human dermal fibroblast NHDF CC-2511 เมื่อทำการทดสอบโดยวิธี MTT reduction assay พบว่าค่า IC50 ของ lipoic acid มีความแตกต่างกันไปในเซลล์เพาะเลี้ยงแต่ละชนิด โดยค่า IC50 ในการยับยั้ง HSV-1 เท่ากับ 126.l85, 109.61 และ 58.20 ug/ml และในการยับยั้ง HSV-2 เท่ากับ 135.06, 125.90 และ 92.26 ug/ml ใน Vero, HeLa และ NHDF CC-2511 ตามลำดับ การศึกษานี้แสดงให้เห็นว่า lipoic acid และ lipoamide มีฤทธิ์ต้านไวรัสเฮอร์ปีส์ซิมเพล็กซ์และอาจนำไปพัฒนาเป็นยารักษาโรคที่เกิดจากการติดเชื้อ HSV ได้ในอนาคต
DOI: 10.14457/cu.the.2007.1608
title: Enhancement of lipase activity from Stenotrophomonas maltophilia by random mutagenesis

description: The isolation of lipase-producing bacteria were isolated from sources of 12 soil samples and 10 composts in Thailand, using enrichment media containing olive oil. The primary screening was carried out employing tributyrin agar plate and Rhodamine B supplemented with olive oil was used for secondary lipase screening. Fifty-five lipase-producing isolates were determined lipase activity by using p-nitrophenyl palmitate as substrate. The results showed that Stenotrophomonas maltophilia CU22 exhibited the highest specific activity. Also, lipase from S. maltophilia was catalyzed transesterification of the palm oil and methanol in non-solvent media and showed 7% methyl ester conversion. To improve lipase activity, random mutagensis by UV irradiation was used. At 10 sec expose time, the death rate with 96.67%, 47 mutants was as subject to test hydrolytic assay. Ten mutants with higher lipase specific activity were subcultured twenty times and then investigated transesterification. UV107, UV1016 and UV 1048 showed %conversion higher than wild type, were 7.56, 9.30 and 7.3 %. DNA and amino acid sequencing of partial lipase gene of wild type and UV1016 , were compared. The result showed nucleotide substitution at 1765, 1766 and 1768 with different amino acid substitution, were obtained. There are Asn568-&gt;Asp, Gln589-&gt;Trp and Arg590-&gt;Gly.
DOI: 10.14457/cu.the.2007.1609
title: Bioactive compounds from sponge-derived fungus strain CRI 247-01 and soft coral-derived fungus strain CRIF1

description: Chemical constituents of two marine-derived fungi, strains CRI 247-01 and CRIF1, were investigated. The sponge-derived fungus strain CRI 247-01 gave a new compound 2, along with two known compounds, 6-methoxymellein (compound 1) and vermillhotin (compound 3). While, a new compound 4 was isolated from the strain CRIF1 which was isolated from a soft coral. Moreover, derivatives of compound 4, methylester (compound 5), benzylester (compound 6), and hydrogenated product (compound 7) were prepared. Chemical structures of these compounds were elucidated on the basis of their spectroscopic data. In this study, compounds 3, 5, and 6 exhibited potent cytotoxic activity against cancer cell lines. Moreover, compound 6 also inhibited antioxidant activity (HL-60 Antioxidation)
DOI: 10.14457/cu.the.2007.1610
title: Poly(methyl methacrylate) and glycidyl-functionalized poly(methyl methacrylate) nano-size latex particles via differential microemulsion polymerization

description: The differential microemulsion polymerization technique was used to synthesize nanoparticles of glycidyl-functionalized poly(methyl methacrylate), PMMA, via a two-step process, by which the amount of sodium dodecyl sulfate (SDS) surfactant required was 1/217 of the monomer amount by weight and the surfactant/water ratio could be as low as to 1/600. The first step was to synthesize the core PMMA nanoparticles having high molecular weight of 10[superscript 6] g mol[superscript -1] using the AIBN oil soluble initiator at 70 [degrees Celsius] for 90 min by drop-wise addition of the monomer. The high molecular weight PMMA latex nanoparticles (~ 20 nm in size) have a rich syndiotactic configuration (53-57 % rr triads). The rate of polymerization increased with an increase in the concentration of the SDS surfactant. The maximum conversion of polymerization was observed at a polymerization time of 3600 s. The nearly constant value of PDI over the whole range of the polymerization time could be attributed to the significance of particle nucleation occurring via a heterogeneous nucleation mechanism. The high molecular weight PMMA nanoparticles have spherical shape with a T[subscript g] of about 125 [degrees Celsius]. The second step was to add glycidyl methacrylate in the PMMA latex via a similar manner to obtain the glycidyl-functionalized PMMA nanoparticles. A core/shell structure of the glycidyl-functionalized PMMA latex nanoparticles observed are composed of a high molecular weight PMMA core with the random copolymer of poly[(methyl methacrylate)-ran-(glycidyl methacrylate)] as a shell layer on the surface. Particle sizes of about 50 nm were achieved and the molecular weight of glycidyl-functionalized PMMA was in the range of about 1x10[superscript 6] to 3x10[superscript 6] g mol[superscript -1]. Approximately 1-2 wt% of glycidyl functional groups were determined using a titration method. The low content detected was due to precipitation and a drying effect. The glycidyl-functionalized PMMA has two regions of T[subscript g] at 90 [degrees Celsius] and 125 [degrees Celsius], which were referred to as poly[(methyl methacrylate)-ran-(glycidyl methacrylate)] and PMMA, respectively. It is very interesting to note that the polymerization conversion reaches a maximum, within a short reaction time in the presence of less surfactant and without the need of a co-surfactant along with a high monomer-to-water ratio.
DOI: 10.14457/cu.the.2007.1611
title: Development of benzoxazine-based resins for wood composite application

description: The curing reaction of benzoxazine resin, properties of benzoxazine alloys, and characteristics of wood composites used benzoxazine-epoxy alloy matrices were investigated. Kissinger, Ozawa, Friedman, and Flynn-Wall-Ozawa methods were utilized to determine the kinetic parameters of the curing reaction of the arylamine-based polyfunctional benzoxazine resins. BA-a resin shows only one dominant autocatalytic curing process with the average activation energy of 81-85 kJ/mol, whereas BA-35x exhibits two dominant curing processes signified by the clear split of the curing exotherms. The average activation energies of low-temperature curing (reaction (1)) and high-temperature curing (reaction (2)) were found to be 81-87 and 111-113 kJ/mol, respectively. The reaction (1) is found to be autocatalytic in nature, while the reaction (2) exhibits nth-order curing kinetics. To modify polybenzoxazine properties, effects of a monofunctional benzoxazine diluent (Ph-a) on properties of a bifunctional benzoxazine resin (BA-a) have been studied. The BA-a/Ph-a mixtures are miscible in nature rendering the properties highly dependent on their compositions. The viscosity of the BA-a resin can be reduced to one third using only about 10% by weight the Ph-a diluent. The addition of the Ph-a resin into the BA-a resin can also lower the liquefying temperature of the resin mixtures whereas the gel point is marginally decreased. The gel point, which depends on the BA-a/Ph-a mixtures and the cure temperature, was determined by the frequency independence of loss tangent in the vicinity of the sol-gel transition. The relaxation exponent values of the copolymer were found to be 0.24-0.55, which is dependent on the cure temperature. Gel time of the BA-a/Ph-a systems decreases with increasing temperature according to an Arrhenius relation with activation energy of 60.6 1.5 kJ mol?1. Flexural moduli of the BA-a/Ph-a polymers also increase with the Ph-a mass fraction, however, with the sacrifice of their flexural strength and glass-transition temperature. To systematically investigate the effect of benzoxazine alloy compositions on the interfacial interaction with Hevea braziliensis woodflour filler, wood-substituted composites from highly filled polybenzoxazine-epoxy alloys were investigated. The reinforcing effect of the woodflour shows the enhancement on the stiffness compared to that of the ternary matrices. The outstanding compatibility between woodflour and polybezoxazine matrix can improve the modulus and thermal curability i.e., glass transition temperature of the wood composites with polybenzoxazine fraction increased.