Rapid one-step separation and purification of recombinant phenylalanine dehydrogenase in aqueous two-phase systems     

Shahbaz Mohammadi H  Omidinia E  Taherkhani H 《Iranian Biomedical Journal》2008,12(2):115-122

Background: Phenylalanine dehydrogenase (PheDH; EC 1.4.1.20) is a NAD+-dependent enzyme that performs the reversible oxidative deamination of L-phenylalanine to phenylpyruvate. It plays an important role in detection and screening of phenylketonuria (PKU) diseases and production of chiral intermediates as well. The main goal of this study was to find a simple and rapid alternative method for purifying PheDH. Methods: The purification of recombinant Bacillus sphaericus PheDH was investigated in polyethylene glycol (PEG) and ammonium sulfate aqueous two-phase systems (ATPS). The influences of system parameters including PEG molecular weight and concentration, pH and (NH4)2SO4 concentration on enzyme partitioning were also studied. The purity of enzyme was analyzed by sodium dodecyl sulfate polyacrylamide gel electrophoresis. Results: A single extraction process was developed for separation and purification of recombinant PheDH from E. coli BL21 (DE3). The optimized conditions for partitioning and purification of PheDH were 9% (w/w) PEG-6,000 and 16% (w/w) (NH4)2SO4 at pH 8.0. The partition coefficient, recovery, yield, purification factor and specific activity values were achieved 58.7, 135%, 94.42%, 491.93 and 9828.88 U/mg, respectively. Also, the Km values for L-phenylalanine and NAD+ in oxidative deamination were 0.21 and 0.13 mM, respectively. Conclusion: The data presented in this paper demonstrated the potential of ATPS as a versatile and scaleable process for downstream processing of recombinant PheDH.  相似文献   

Inotropic and chronotropic effects of 6-hydroxy-4-methylquinolin-2(1H)-one derivatives in isolated rat atria     

Mansouri SM  Shafiee-Nick R  Parsaee H  Seyedi SM  Saberi MR  Sadeghian H 《Iranian Biomedical Journal》2008,12(2):77-84

Background: Selective phosphodiesterase (PDE3) inhibitors improve cardiac contractility and may use in congestive heart failure. However, their proarrhythmic potential is the most important side effect. Methods: In this research, we evaluated the potential cardiotonic activity of six new synthesized selective PDE3 inhibitors (6-hydroxy-4-methylquinolin-2(1H)-one derivatives) using the spontaneously beating atria model. In each experiment, atrium of reserpine-treated rat was isolated and the contractile and chronotropic effects of a synthesized compound were assessed. The 3-isobutyl-1-methylxanthine, a non-selective PDE inhibitor, was used for comparison. Results: The results showed that, among new compounds, the best pharmacological profile was obtained with the compound 6-[4-(4-methylpiperazine-1-yl)-4-oxobutoxy]-4-methylquinolin-2(1H)-one, C6, which displayed selectivity for increasing the force of contraction (168 +/- 5% change over the control) rather than the frequency rate (138 +/- 5% change over the control) at 300 muM. However, C6 at concentrations of 10 and 100 muM produced left and upward shift in the positive inotropic concentration-response curve of isoprenaline. The -log EC50 of isoprenaline was 8.843 +/- 0.171 in the absence, 9.448 +/- 0.138 and 9.456 +/- 0.107 in the presence of 10, 100 muM of C6, respectively (P<0.001, n = 6). Also, amrinone, a selective PDE3 inhibitor, shifted the isoprenaline concentration-response curve to the left and upward. The concentration of 10 and 100 muM amrinone decreased -log EC50 of isoprenaline to 9.527 +/- 0.287 and 9.423 +/- 0.243, respectively (P<0.001, n = 6). Moreover, the positive chronotropic effect of isoprenaline was not affected by amrinone or C6. Conclusion: This study provides functional evidence for the positive inotropic effect of C6. Considering the augmentation of isoprenaline positive inotropic concentration-response with C6 and amrinone, we conclude that C6 produces its effect via potentiation of cAMP-dependent signaling system and possibly by inhibition of PDE3 activity.  相似文献   

Beta cell protective effects of sodium tungstate in streptozotocin-induced diabetic rats: glycemic control, blockage of oxidative stress and beta cell histochemistry     

Heidari Z  Harati M  Mahmoudzadeh-Sagheb HR  Moudi B 《Iranian Biomedical Journal》2008,12(3):143-152

Background: Diabetes is a major public health problem. The development of new therapies that are able to improve glycemia management and even to cure diabetes is of great interest. In this study, protective effects of sodium tungstate against STZ-induced beta-cell damages were investigated. Methods: Sixty rats were divided into six groups: control, diabetic, sodium tungstate treated diabetic rats from one week before STZ injection (TDB), food-restricted diabetic (FRD), tungstate treated control, sodium tungstate treated diabetic rats from one week after STZ administration (TDA). We evaluated serum insulin, glucose and glucose tolerance; liver glycogen content, glucokinase (GK) activity; blood and pancreas antioxidant power, lipid peroxidation; and fuchsin-aldehyde histochemical staining of beta-cells. Results: Blood glucose levels of TDB group were lower than other diabetic groups (P<0.01). Blood insulin levels of all diabetic groups were lower than controls (P<0.01). Glucose intolerance improved in TDB animals. Blood and pancreas antioxidant power, liver glycogen contents and GK activities and granulated beta cells increased in TDB rats in comparison with other diabetic groups (P<0.01). Likewise, lipid peroxidation decreased significantly in TDB rats (P<0.01). Conclusions: Results suggested that sodium tungstate if administrated before STZ injection improves glycemic state by a direct effect on pancreatic beta-cells and preserves them by reducing the activity of these cells at the time of STZ injection, reducing STZ-induced oxidative stress, reducing insulin secretion, or all of the above mentioned.  相似文献   

New phenantrene alkaloids from Cryptocarya crassinervia   总被引：1，自引：0，他引：1  

Awang K  Hadi AH  Saidi N  Mukhtar MR  Morita H  Litaudon M 《Fitoterapia》2008,79(4):308-310

The bark of Cryptocarya crassinervia provided two new phenantrene alkaloids, 2-hydroxyatherosperminine (1) and N-demethyl-2-methoxyatherosperminine (2).  相似文献   

Alleviation of dormancy in walnut kernels by moist chilling is independent from storage protein mobilization   总被引：1，自引：0，他引：1  

Einali AR  Sadeghipour HR 《Tree physiology》2007,27(4):519-525

We studied the effects of moist chilling and warming on storage protein mobilization in walnut (Juglans regia L.) kernels to assess the metabolic inhibition theory, which states that dormant seeds are unable to utilize their own food reserves and that cold conditions allow germination by activating hydrolases involved in reserve mobilization. Stratifying kernels at 5 degrees C for 40 days enhanced their germination. During both cold stratification and warm incubation of kernels, storage protein mobilization occurred in cotyledons rather than axes. Kernel amino acid concentration increased during protein mobilization, with axes of warm-incubated kernels having particularly high amino acid concentrations. Polyacrylamide gel electrophoresis in the presence of sodium dodecyl sulfate (SDS-PAGE) of the soluble protein fractions from both cold-stratified and warm-incubated cotyledons revealed increased band intensities of putative glutelins (19-22 and 32-35 kDa). A very high molecular weight protease was detected by gelatin SDS-PAGE that was most active at acid to neutral pHs in imbibed, cold stratified and germinated kernels suggesting the protease(s) was synthesized earlier in the mature seeds. Thus, in dormant walnut kernels there is no block to protein mobilization, and imbibition alone is sufficient to initiate proteolysis. Catalase activity was higher in warm-incubated kernels than in cold-stratified kernels, suggesting that seed aging is hastened under warm conditions and that cold stratification in walnut kernels might involve activation of cellular repair mechanisms.  相似文献   

Potassium Fixation and Release in Some Calcareous Soils under Orange Cultivation     

Mahdi Najafi-Ghiri  Boostani  Hamid Reza  Mirsoleimani  Abbas  Mohaviye-Asadi  Negin  Beizavi  Majid  Shafiei  Mina  Mirdoraghi  Maryam 《Eurasian Soil Science》2020,53(7):978-985

Eurasian Soil Science - Arid and semiarid soils of southern Iran may fix a large content of applied potassium (K), but the fixed K may be gradually supplied to plants. Sixteen representative soils...  相似文献   

Effect of zinc bacitracin and phytase on growth performance,nutrient digestibility,carcass and meat traits of broilers          下载免费PDF全文

Y. A. Attia  F. Bovera  A. E. Abd El‐Hamid  A. E. Tag El‐Din  M. A. Al‐Harthi  A. S. El‐Shafy 《Journal of animal physiology and animal nutrition》2016,100(3):485-491

A total of 336 one‐day‐old Hubbard broiler chickens were randomly distributed among 8 groups, each containing six replicates (7 chickens/replicate). From 1 to 40 days of age, the groups fed the same starter, grower and finisher diets. The control group was unsupplemented; zinc bacitracin (ZnB) group received the antibiotic at 0.5 g/kg; fungal phytase (FP) groups received 250, 500 and 1000 U/kg diet of Aspergillus niger phytase (FP_250, FP_500 and FP_1000 groups), respectively; bacterial phyatse (BP) groups received 250, 500 and 1000 U/kg diet of Escherichia coli phytase (BP_250EP, BP_500EP and BP_1000EP groups) respectively. Considering the whole experimental period, body weight gain was unaffected by ZnB and different concentrations of bacterial and fungal phytase; however, the feed conversion ratio of the group fed a diet supplemented with 500 U of BP was better (p < 0.01) than those fed with a diet supplemented with 500 U of FP. BP_250 group had a higher (p < 0.05) apparent digestibility of ether extract compared to FP_250 group. In conclusion, bacterial phytase at 500 U may enhance performance of broiler chickens fed during days 1–40 of age and yield similar growth performance and economic efficiency to those of eB‐supplemented groups.  相似文献   

Effects of hypertonic dextrose and paraffin solution as non‐antibiotic treatments of clinical endometritis on reproductive performance of high producing dairy cows     

Mohammad Rahim Ahmadi  Meysam Makki  Abdolah Mirzaei  Hamid Reza Gheisari 《Reproduction in domestic animals》2019,54(5):762-771

The objective of this study was to compare the reproductive performance of cows affected by clinical endometritis (CE) following treatment with an intrauterine infusion of 50% dextrose solutions (DEX) and liquid paraffin (LP) as alternative therapies with routine treatments including PGF_2α injection and intrauterine infusion of oxytetracycline (OTC). Moreover, the reproductive indices of cows treated with endometritis were compared with those of healthy ones. At postpartum reproductive tract examination (28–35 DIM) in one Iranian dairy farm, cows with CE without any selection were assigned to four groups: (a) OTC, a common treatment in Iran, was administered (5 g) to 396 cows, (b) PGF₂α (PG) was injected to 496 cows, (c) dextrose solution (DEX): intrauterine infusion of 50% dextrose solution (200 ml) was done in 427 cows, and (d) liquid paraffin (LP) was administered (100 ml) to 423 cows via intrauterine route. We further assigned 2,233 clinically healthy cows to the control group. The incidence of endometritis was 41.6% in this study. Based on the results of reproductive indices including median days to first AI, days open (DO), first service conception rate, conception rate in 2nd and 3rd services, conception rate in all three services, pregnancy rate < 100 days and pregnancy rate < 200 days, except for median days to first AI in other reproductive indexes, reproductive performance was significantly lower in LP group compared with the healthy cows and other treatment groups (p < 0.05). Except for the first service conception rate and proportion of cows pregnant before 100 days in milk (DIM), there existed no significant difference between the DEX group and the control as far as reproductive performance is concerned (p ≥ 0.05). The first service conception rate was recognizably lower in DEX group compared with OTC and PG (p < 0.05). In conclusion, the use of a non‐antibiotic special solution of dextrose 50% is a good alternative to antibiotic agents concerning the treatment of CE in dairy cows.  相似文献   

Effect of adding different levels of caffeine in the extender on some biochemical constituents,enzymatic activities and physical characteristics of chilled and frozen ram semen     

Ibrahim S. Abd El‐Hamid 《Reproduction in domestic animals》2019,54(2):225-233

Two experiments were carried out to determine the efficiency of supplementation of ram semen extender with caffeine on chilled storage and frozen capacity of spermatozoa. In the first experiment, eighty ejaculates were collected by an artificial vagina from five adult Barki rams, aged 2–3 years and weighted 45.0 ± 2.0 kg throughout the experimental period (January to February 2017). The ejaculates were pooled and diluted (1:10) with tris‐citric egg yolk extender and were split into five groups. Group 1 served as control, whereas groups 2‐5 were supplemented with 0.1, 0.2, 0.3 and 0.4 mM caffeine. All diluted semen specimens were evaluated for physical characteristics immediately after dilution (T₀) and throughout preservation period of 48 hr at 4°C. Simultaneously, oxidative stress and indices such as total antioxidant capacity (TAC), malondialdehyde concentrations (MDA) and alkaline transaminase (AKP) concentrations and value of resazurin reduction test (RRT) were determined. In the second experiment, the raw pooled ejaculates were diluted (1:10) with glycerolated tris‐citric egg yolk extender, receiving the previously mentioned caffeine levels. The post‐thaw assessment of cryopreserved spermatozoa, in all groups, was conducted by a computer‐assisted sperm analysis (CASA) system. The results revealed that adding caffeine to ram semen extender at low (0.1 mM) or medium (0.2 mM) levels had positive impact on both physical characteristics of ram sperm and the enzymatic activities compared to the other semen groups. Caffeine supplementation also enhanced post‐thaw sperm dynamics, which implies its potential as an exogenous antioxidant supplement.  相似文献   

Alleviation of mycotoxin biodegradation agent on zearalenone and deoxynivalenol toxicosis in immature gilts     

Donghui Shi  Jianchuan Zhou  Lihong Zhao  Xiaoping Rong  Yu Fan  Humera Hamid  Wenqiang Li  Cheng Ji  Qiugang Ma 《畜牧与生物技术杂志(英文版)》2018,(3)

Background: The current study was carried out to evaluate the effects of mycotoxin biodegradation agent(MBA, composed of Bacillus subtilis ANSB01 G and Devosia sp. ANSB714) on relieving zearalenone(ZEA) and deoxynivalenol(DON) toxicosis in immature gilts.Methods: A total of forty pre-pubertal female gilts(61.42 ± 1.18 kg) were randomly allocated to four diet treatments: CO(positive control); MO(negative control, ZEA 596.86 μg/kg feed and DON 796 μg/kg feed);COA(CO + 2 g MBA/kg feed); MOA(MO + 2 g MBA/kg feed). Each treatment contained 10 replicates with 1 gilt per replicate. Gilts were housed in an environmentally controlled room with the partially slatted floor.Results: During the entire experimental period of 28 d, average daily gain(ADG) and average daily feed intake(ADFI)of gilts in MO group was significantly reduced compared with those in CO group. The vulva size of gilts was significantly higher in MO group than CO group. In addition, significant increases in the plasma levels of Ig A,Ig G, IL-8, IL-10 and PRL were determined in MO group compared with that in CO group. ZEA and DON in the diet upregulated apoptotic caspase-3 in ovaries and uteri, along with down-regulated the anti-apoptotic protein Bcl-2 in ovaries. The supplementation of MBA into diets co-contaminated with ZEA and DON significantly increased ADG, decreased the vulva sizes, reduced the levels of Ig G, IL-8 and PRL in plasma, and regulated apoptosis in ovaries and uteri of gilts.Conclusions: The present results indicated that feeding diet contaminated with ZEA and DON simultaneously(596.86 μg/kg + 796 μg/kg) had detrimental effects on growth performance, plasma immune function and reproductive status of gilts. And MBA could reduce the negative impacts of these two toxins, believed as a promising feed additive for mitigating toxicosis of ZEA and DON at low levels in gilts.  相似文献