Using recombinant human growth hormone as a model protein, we carried out unfolding by adding a denaturant such as urea, guanidine HCl, or SDS followed by refolding by dilution and dialysis. The objectives were to monitor the structural changes during in vitro refolding process and, based on the results, to develop a quantitative method of refolding progress assessment. The changes in surface hydrophobicity were measured by fluorescence tagging of 1-anilinonaphthalene-8-sulfonate(1,8-ANS) to the hydrophobic portions, and those in the secondary structure were monitored by using far UV-CD(circular dichroism) spectroscopy. Also, we used RP-HPLC to separate and quantify the folded and unfolded proteins to correlate the result with the structure analysis. Our results indicate the surface hydrophobicity are well correlated with the formations of the secondary structure, primarily ${\alpha}$-helices, as well as the disulfide bridges. We expect this monitoring technique can be applied in industrial fields as a means to quantitatively assess the progress of in-vitro refolding of recombinant proteins.
Surfactant-enhanced air sparging (SEAS) was developed to suppress the surface tension of groundwater prior to air sparging resulting in higher air saturation and larger contact area between NAPL and gas during air sparging. Larger contacting interface between NAPL and gas means faster mass transfer of contaminants from NAPL to gas phase. This new technique, however, is limited to relatively volatile contaminants because vaporization is its basic mechanism of mass transfer. In this study, SEAS was tested at an elevated temperature for a semi-volatile n-decane, which is expected not to be a good candidate of SEAS application due to its low vapor pressure at ambient temperature. Three sparging experiments were conducted using 1-dimensional column (5 cm id, 80 cm length) packed with sand; (1) ambient temperature ($23^{\circ}C$), column saturated with distilled water, (2) SEAS at ambient temperature ($23^{\circ}C$), for n-decane contaminated sand, (3) SEAS at elevated temperature ($73^{\circ}C$), for n-decane contaminated sand. Higher air saturation was achieved by SEAS compared to that by air sparging without surfactant application. The n-decane removal efficiency of SEAS at elevated temperature was significantly higher(> 10 times) than that of ambient SEAS. The n-decane concentrations in the gas effluent from column during SEAS at $73^{\circ}C$ are found to be 10 times of those measured at ambient temperature. Thus, SEAS technique can be applied for removal of semi-volatile contaminants provided that an appropriate technique for elevating aquifer temperature is available.
Park, Chang-Mo;Kim, Tae-Young;Kim, Won-Jung;Kim, Yun-Sang;Suh, Kwang-S
Polymer(Korea)
/
v.29
no.4
/
pp.363-367
/
2005
The effects of organic solvent on the charge transport behavior of poly (3,4-ethylenedioxythioph one)/p-toluene-sulfonate(PEDOT-OTs) are investigated. The use of different organic solvents during the oxidative chemical polymerization of 3,4-ethylenedioxythiophene (EDOT) with Iron(III) -tosylate can greatly vary the DC conductivity of PEDOT-OTs along with molecular structure and doping concentration. For example, PEDOT-OTs prepared from methanol shows the conductivity of 19.5 S/cm, which is an increase by a factor of $10^8$ compared to PEDOT-OTa prepared from acetone. From the X-ray diffraction (XRD) experiments, it was found that PEDOT-OTs with ketone is amorphous state, while PEDOT-OTs with alcoholic solvent shows the better defined crystalline structure in which the charge transport along and between the PEDOT chains are promoted. Chemical analysis employing X-ray photoelectron spectroscopy (XPS) revealed that the doping concentration of PEDOT-OTs with alcoholic solvent is much higher than that of PEDOT-OTs with ketones. It is proposed that the interactions between the organic solvent and doping anion can cause the variation in doping concentration and, therefore, result in the PEDOT-OTs of different conductivities and chain structures.
Bacillus clausii I-52 which produced SDS- and $H_2O_2$-tolerant extracellular alkaline protease (BCAP) was isolated from heavily polluted tidal mud flat of West Sea in Incheon, Korea and stable strain (transformant C5) of B. clausii I-52 harboring another copy of BCAP gene in the chromosome was developed using the chromosome integration vector, pHPS9-fuBCAP. When investigated the production of BCAP using B. clausii transformant C5 through pilot-scale submerged fermentation (500 L) at $37^{\circ}C$ for 30 h with an aeration rate of 1 vvm and agitation rate of 250 rpm, protease yield of approximately 105,700 U/mL was achieved using an optimized medium (soybean meal 2%, wheat flour 1%, sodium citrate 0.5%, $K_2HPO_4$ 0.4%, $Na_2HPO_4$ 0.1%, NaCl 0.4%, $MgSO_4{\cdot}7H_2O$ 0.01%, $FeSO_4{\cdot}7H_2O$ 0.05%, liquid maltose 2.5%, $Na_2CO_3$ 0.6%). The enzyme stability of BCAP was increased by addition of polyols (10%, v/v) and also, the stabilities of BCAP towards not only the thermal-induced inactivation at $50^{\circ}C$ but also the SDS and $H_2O_2$-induced inactivation at $50^{\circ}C$ were enhanced. Among the polyols examined, the best result was obtained with propylene glycol (10%, v/v). The BCAP supplemented with propylene glycol exhibited extreme stability against not only the detergent components such as ${\alpha}$-orephin sulfonate (AOS) and zeolite but also the commercial detergent preparations. The granulized enzyme of BCAP was prepared with approximately 1,310,000 U/g of granule. Wash performance analysis using EMPA test fabrics revealed that BCAP granule exhibited high efficiency for removal of protein stains in the presence of anionic surfactants as well as bleaching agents. When compared to Savinase 6T$^{(R)}$ and Everlase 6T$^{(R)}$ manufactured by Novozymes, BCAP under this study probably showed similar or higher efficiency for the removal of protein stains. These results suggest that the alkaline protease produced from B. clausii transformant C5 showing high stability against detergents and high wash performance has significant potential and a promising candidate for use as a detergent additive.
Journal of Korean Society of Environmental Engineers
/
v.36
no.2
/
pp.119-126
/
2014
The object of this study was to evaluate the combined toxic interactions of the perfluorooctanoic acid (PFOA) or perfluorooctane sulfonate (PFOS) with six heavy metals (Cu, Zn, Cr, Cd, Hg, and Pb). The individual and combined toxic effects were assessed using the Vibrio fischeri assay. In case of the individual toxicity, PFOA was higher toxic than PFOS and toxicity of PFOA and PFOS were lower than heavy metal. In the toxicity of heavy metals, the $Hg^{2+}$ was found to be most toxic followed by $Pb^{2+}$, $Cr^{6+}$, $Cu^{2+}$, $Zn^{2+}$, and $Cd^{2+}$. The combined toxicity of PFOA or PFOS with $Cr^{6+}$ were synergistic effect because the $EC_{50}$ mix values were less than 1 TU. PFOA + $Zn^{2+}$, PFOS + $Zn^{2+}$, PFOA + $Cd^{2+}$ and PFOS + $Cd^{2+}$ produced addictive effect. Except in these case, all of binary mixtures show antagonistic effect. This study proved potential risk of coexistent with perfluorinated compounds and heavy metals in water environment.
Ayisi, Christian Larbi;Zhao, Jinliang;Yame, Chen;Apraku, Andrews;Debra, Grace
Fisheries and Aquatic Sciences
/
v.22
no.11
/
pp.25.1-25.9
/
2019
Background: Due to the continuous demand for fish coupled with decline in capture fisheries, there is the need to increase aquaculture production to meet the demand. Aquaculture is faced with high cost of feeding since fish oil and fish meal are expensive. In view of this, there are calls to explore alternatives that are cheap and reliable. Objectives: This study on Oreochromis niloticus was conducted to evaluate the effects of replacing fish oil (FO) with palm oil (PO) at 0%, 25%, 50%, 75%, and 100% on muscle fatty acid and proximate composition as well as growthrelated enzyme activities and mRNA expression. Methods: Oreochromis niloticus were fed five experimental diets (33% crude protein and 10% crude lipid) for 8 weeks. Feed had variation in fish oil and palm oil contents. After the 8 weeks feeding trial, five fish were sampled from each tank (15 from each treatment) and euthanized using an excess dose of tricaine methane sulfonate (MS-222 at 200 mg/L). Fatty acid and enzyme activities were analyzed using standard protocols. Also, RT-qPCR was used to quantify the expression levels of selected growth-related genes. Results: Fish fed 25% PO recorded the least muscle protein content and was significantly lower than the group fed 100% PO. Paired box protein 7 (Pax-7) enzyme activity was significantly higher in the group fed 50% PO compared to the groups fed 25% PO and 100% PO, while caplain-3 (Capn-3) was significantly lower in the group fed 0% PO compared to all other groups. There was a significant difference among treatments with respect to mRNA expression of Pax-7 and Capn-3. Group fed 25% PO had significantly lower mRNA expression of Pax-7, while the group fed 75% PO recorded significantly higher mRNA expression of Capn-3 compared to groups fed 0% PO, 25% PO, and 100% PO. Pearson's correlation analysis revealed that Igf-I and Igf-II mRNA expression have significant correlation with n-3 polyunsaturated fatty acids content in muscle. Conclusion: The results suggest muscle protein content could be modified if FO is replaced with PO. Also, mRNA expression of Pax-7 and Capn-3 is affected by replacing FO with PO.
In this study, a sulfonated poly(arylene ether ketone) block copolymer was prepared from hydrophilic oligomer and hydrophobic oligomer. The structure of the prepared membrane was characterized by $^1H$-NMR, FT-IR and GPC. The $M_w$(weight-average molecular weights) of the polymer was $209,700g\;mol^{-1}$ and the molecular weight distribution($M_w/M_n$) of 1.25 was obtained. The prepared membrane showed excellent thermal stability with gradual weight loss up to $200^{\circ}C$. The proton conductivity of SPAEK block copolymer reached the maximum of $9.0mS\;cm^{-1}$ at $90^{\circ}C$ under 100% relative humidity (RH). From the observed results, it is necessary to do more aggressive attempt to study the possibility of application as an ion-conductive composite electrolyte.
This experiment was conducted to find out selective herbicides which are safe to the rice seedlings and to provide effective weed control method in protected semi-irrigated rice seedbed. There was no crop injury in rice with benzophenap [2-(4-2, 4-dichloro-3-methylbenzoyl)-1, 3-dimethyl-pyrazol-5-yl-oxy)-4'-methyl acetophenone] (240g), pyrazoxyfene [1,3-dimethyl-4-(2,4-dichlorobenzoyl)-Sphenacyloxy pyrazole] (200g), chlormethoxynil [2,4-dichlorophenyl-4-nitro-3-methoxyphenyl ether] (180g), dimepiperate [S-(1-methyl-1-phenethyl)-piperidine-l-carbathioate] (210g), dimepiperate + probenazol [3-allyloxy-1,2-benzisothiazol-1,1-dioxide] (210 + 120g) mixture and dimepiperate + probenazol + molinate [S-ethyl-N,N-hexamethylene-thiol-carbamate] (120 + 120 + 120g) mixture at times of application studied. Butachlor [2-chloro-2',6'-diethyl-N-buthoxymethyl acetanilide] + pyrazolate [4-(2,4-dichlorbenzoyl)-1,3dimethyl-pyrazol-5-yl-p-toluene-sulfonate] (70 + 120g) mixture and butachlor + chlormethoxynil (60 + 120g) mixture caused root length to shorten and root viability to decrease. However, the crop injury was recovered at 25 days after seeding. Benzophenap, pyraxoxyfene and butachlor + pyrazolate mixture were effective for weed control when applied at 2 days before seeding. Chlormethoxynil and butachlor + chlormethoxynil mixture was very effective for controlling annual weed. Dimepiperate, dimepiperate + probenazol mixture and dimepiperate + probenazol + molinate mixture did not control most weeds except for Echinochloa crus-galli (L.) Beauv.
In order to produce xylitol with high yield, experiments were carried out to develope xylitol dehydrogenase(XDH) defective mutant from P. stipitis and to investigate the xylitol fermentation characteristics of mutant strain. After treatment of P. stipitis with EMS, mutant PXM-4 was selected based on te XDH activity and xylitol production capability. Among the tested cosubstrates, galactose was selected as an adequate cosub-strate on xylitol production of mutant PXM-4. But with the increase in the concentration of galactose in the medium, xylitol production was decreased because the transport of xylose into cell was inhibited by galactose. The optimal concentration of galactose for the production of xylitol using 20 g/ι xylose was 20 g/ι Under this condition, maximum concentration of xylitol and yield were 14.4 g/ι and 97%, respectively. In order to prevent the inhibitory effect of xylose transport by galactose, galactose was fed with low concentration and the concentration of xylitol produced was increased up to 25 g/ι.
It has been reported from our department that a few agents, such as $K_2S_2O_5,\;NaHSO_3,$ nicotinamide have a marked stabilizing effect in vitro on metoclopramide which is relatively unstable compound. In order to study the effect of these stabilizers on the action of metoclopramide in vitro, the fate of this compound combined with $K_2S_2O_5,\;NaHSO_3$ and nicotinamide, respectively, was studied and furthermore, the change of the biological activity of metoclopramide due to these stabilizers was studied by using the isolated stomach strip of rat. The blood concentration of metoclopramide was measured by using Bakke's method at the various time after intravenous injection of the mixed metoclopramide solution with the stabilizers. In order to study the excretion of the drug, rabbits were anesthesized and catheterized into bladder for withdrawal of urine. After intravenous injection of the mixed metoclopramide solution, urine was collected for 5 hours and the conjugated forms of metoclopramide as well as the free form were determined by using Arita's method. In the biological study of the metoclopramide combined with stabilizers, the contractability of the isolated rat stomach strip was observed by using polygraph recorder. The results were following: 1. When metoclopramide was administered with nicotinamide as stabilizer, the blood concentration of the unchanged from and the rate of the clearance of this compound were very similar to that of metoclopramide alone. On the other hand, other stabilizers, $K_2S_2O_5\;and\;NaHSO_3$, brought about 40% decrease in blood concentration of the unchanged form at 15 min after intravenous injection however, the rate of clearance of metoclopramide with $K_2S_2O_5\;or\;NaHSO_3$ was very slow. 2. In the case of urinary excretion, the excretory pattern of the metabolites of metoclopramide with $NaHSO_3$ or nicotinamide was very similar to that of metoclopramide alone. But metodopramide plus $K_2S_2O_5$ group showed the maked depression of excretion for first 1 hour. 3. In composition of metabolites, when metoclopramide was administered with $K_2S_2O_5$ or $NaHSO_3$, the sulfonate conjugation was predominant. But the glucuronic acid conjugation was predominant in metoclopramide plus nicotinamide gronp. 4. In the experiments on the biological activity of the metoclopramide, this compound exhibited the marked contracting effect in isolatd rat stomach strip. Specially, the meetoclopramide combined with $K_2S_2O_5$ showed the strong contraction of the isolated strip, suggesting the potenciating effect of $K_2S_2O_5$ on the action of metoclopramide in the isolated strip.
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