Ochrobactrum intermedium DN2烟碱降解酶发酵培养基优化研究

研究工作中筛选得到一株有较强烟碱降解能力的新型烟碱降解细菌 Ochrobactrum intermedium DN2,对其产烟碱降解酶的发酵培养基进行了优化.采用Plaekett-Burman设计法,对影响Ochrobactrum intermedium DN2产烟碱降解酶的15个因子进行了筛选.结果表明,影响该菌发酵产酶的显著因子为烟碱、酵母膏、葡萄糖、tween-80的质量浓度和培养基初始pH.在此基础上,采用响应曲面法对以上5个显著因子进行了优化.得到各因子最佳水平为烟碱2.183g/L,葡萄糖0.823 g/L,酵母膏0.844 g/L,Tween-80 0.976 g/L,初始pH 6.9.在此优化条件下获得实际酶活为7943 U/L,与预测值8 060 U/L相近,比优化前提高了52%.     

利用响应面分析法优化γ-氨基丁酸发酵培养基

通过响应面分析的方法对发酵生产γ 氨基丁酸(GABA)的培养基进行优化.利用二水平正交试验考察葡萄糖、豆饼粉、玉米浆、K2HPO4、吐温 80、起始pH值和谷氨酸钠(MSG)对发酵生产GABA的影响.利用极差分析找出主要影响因子:分别为豆饼粉、玉米浆和葡萄糖.利用中心组合设计与响应面分析进一步考察主要影响因子并确定了最佳培养基的组成.在优化培养基中,GABA产量增加约4倍,达到3.63g/L,实验值与预测值基本相符.     

植物乳杆菌HO-69产抗菌肽的培养条件

对L．plantarum HO-69产抗菌肽的营养和培养条件进行了研究。实验结果袁明，MRS培养基是其产抗菌肽的最适培养体系。以碳源、氮源、缓冲盐等为研究因子，对MRS设计部分因子重复实验，采用回归分析确定K2HPO4和牛肉膏为抗菌肽产生的显著影响因子，综合考虑抗菌肽活力与纯化的难度，确定K2HPO4与牛肉膏的质量浓度分别为12g／L与16g／L，蛋白胨质量浓度为6g／L。响应面分析确定HO-69产抗菌肽的最适培养条件为：起始pH值6．61，36．12℃发酵13．87h，在此条件下发酵液的效价由80AU／mL提高到320AU／mL，抗菌肽的产量增加为原来的4倍。     

Penicillium sp.X-1液态发酵生产生淀粉酶的优化

通过摇瓶发酵,研究了培养基成分对Penicillium sp.X-1液态发酵产生淀粉酶的影响。结果表明：碳源、氮源及MgCl2对产酶有较大的影响,经响应面优化得到的培养基组成为：玉米粉42 g/L,豆饼粉30 g/L,MgCl216 mmol/L,在最优条件下酶活达到239 U/mL,与采用基本培养基的相比,酶活提高了7.5倍.     

正交旋转回归试验优化纤维素酶发酵培养基

利用旋转回归法研究里氏木霉WX-112发酵生产纤维素酶的两个重要因素：微晶纤维素粉（Avicel）和麸皮对滤纸酶活的影响，并拟合出回归方程。经回归分析表明，培养基中Avicel、麸皮的含量及其配比对滤纸酶活有显著影响。通过岭脊分析寻优得出：Avicel最佳浓度为1．34g／dL、麸皮最佳浓度为3．35g／dL，在此优化条件下滤纸酶活可迭6．51U／mL。用30L发酵罐进行放大试验，滤纸酶涪可达10．84U／mL，CMCase达到449．57U／mL。     

阿片肽酿酒酵母工程菌摇瓶发酵工艺

研究了发酵培养基成分、补加方式及外源氨基酸对酿酒酵母发酵和表达的影响．结果表明，发酵前在液体发酵培养基中一次性补加酵母抽提物（YE）有利于目的产物的表达，发酵过程中补加YE不利于目的产物的表达，发酵中后期补加适量的葡萄糖有利于阿片肽的表达．添加外源氨基酸对酿酒酵母工程菌的生长有一定的促进作用，而且可以明显地影响阿片肽的表达．通过正交实验对酿酒酵母生长和表达条件进行优化，确定最优工艺参数为：培养基中酵母抽提物质量浓度为25g／L，C：N比2．8，培养基初始PH5．0，摇床转速220r／min．在最优条件下研究了酿酒酵母表达目的产物的情况，总蛋白质质量浓度为614．50mg／L．SDS-PAGE电泳和双波长凝胶扫描结果表明：目的阿片肽约占总分泌蛋白质的5％，其表达量为30．7mg／L，是优化前的1．49倍。     

基于遗传算法的木糖醇发酵培养基的优化

运用遗传算法，利用莫格假丝酵母由木糖生产木糖醇的发酵培养基进行优化，用40个实验样本完成了6种培养基成分、50个浓度水平的优化任务.实验结果表明利用遗传算法可优化培养基成分含量，取得更好的发酵效果.按照优化后的培养基组成，由50g／L木糖获得了29.7g／L木糖醇，理论转化率为65.1%，比优化前提高了3.5%.     

谷氨酸脱羧酶发酵工艺的优化

用大肠杆菌AS1.505进行液态发酵生产谷氨酸脱羧酶并优化培养基,考察了碳源、氮源、复合营养物质、起始pH及发酵时间对酶活的影响,确定最佳产酶培养基组成为:葡萄糖1.0 g/dL,蛋白胨3.0 g/dL,氯化钠0.3 g/dL,磷酸氢二钾0.1 g/dL,硫酸镁0.02 g/dL,L-谷氨酸0.01g/dL,玉米浆1.5 g/dL,生物素30 t,g/L,麸皮4 g/dL;pH 6.5.在此基础上,设计发酵条件的优化实验.实验结果表明为:250 mL的三角瓶装液量25 mL,37℃,起始pH 6.5,培养18 h达到产酶高峰,产酶活力可达1 290 U/mL.     

Thermobifida fusca WSH03-11突变株合成角质酶的发酵条件

以放线菌Thermobifida fusca WSH03—11——高产角质酶突变株为出发菌株，在摇瓶中考察了碳、氮源等条件以及接种量、装液量和初始pH值等环境条件对突变株细胞生长和产角质酶的影响。通过优化发酵条件，突变菌株的角质酶酶活达10．1U／mL，提高了15％。在此基础上，在5L发酵罐中进一步研究了碳源流加对突变株发酵产酶的影响，结果发现，分别在0、24、48h添加1％乙醇，角质酶酶活达到16．4U／mL，细胞干重（DCW）达到3．7g／L，而且发酵时间也由110h缩短到50h。     

产胆固醇氧化酶重组大肠杆菌的发酵培养基和诱导条件的优化

在摇瓶中对产胆固醇氧化酶重组大肠杆菌的发酵培养基和诱导条件进行了优化,优化培养基为甘油10 g/L,胰蛋白胨10 g/L,酵母粉5 g/L,KH2PO4 2 g/L,K2HPO4 4 g/L,Na2HPO4·12H2O 7 g/L,(NH4)2SO4 1.2 g/L,NH4Cl 0.2 g/L,MgSO4·7H2O 1 g/L;优化的诱导条件为:对数生长中期诱导,IPTG浓度为0.3 mmol/L.在优化的培养基和优化的诱导条件下,单位菌体产酶量达745.86 U/g,菌体产酶水平达1 625.97 U/L,为优化前的700 U/L的2.3倍.     

Validation of the gas exchange monitoring function of the Engstr?m Elvira respirator]

Gas exchange monitoring (GEM) has only recently become available for routine use on ventilated patients. Engstr?m has introduced a new concept of integrated gas exchange monitoring within the Elvira respirator, optionally combined with an external CO2 monitor. The following paper describes a laboratory validation of a prototype of this device with regard to the influence of respiratory variables within their specified ranges: FIO2 0.2-0.6, FIO2-FEO2 (DFO2) 0.02-0.05, FECO2 0.02-0.05, VI 5-20 l/min, p 10-60 mbar, respiratory rate 7-29, VT 500-2,000 ml, inspiratory flow 25-90 l/min, relative humidity 5% and 55%. VO2 and VCO2 (100-700 ml/min) were simulated by gas dilution at a respiratory quotient of 1 using a 'bag-in-the-bottle' test lung for 48 test situations. Two reference measurements per test situation were carried out by mass spectrometry and wet-gas spirometry. The Elvira GEM function uses a self-calibrating fuel cell for FO2 detection, an external infrared CO2 monitor (Eliza) for FECO2 measurement, and differential pressure detection on a venturi principle for inspiratory flow measurement. VI measurement necessitates the use of the Haldane transformation for VO2 and VCO2 calculation. The VO2 (VCO2) repeatability coefficient (2 s) for the reference method was 5.6 (5.5) ml/min compared to 10.3 (10.35) ml/min for the GEM function. The mean relative methodical difference for VO2 (VCO2) was +19% (+4.1%) with limits of agreement (+/- 2 s) of +/- 13% (+/- 8.7%). The systematical difference for VCO2 as well as the variability within different test situations was in an acceptable range for clinical measurement.(ABSTRACT TRUNCATED AT 250 WORDS)     

Gas chromatographic determination of the solubility coefficient for volatile anaesthetics (author's transl)]

For the determination of the solubility coefficient of volatile anaesthetics in Schindler's liquid culture, modified by Karzel, a gas-chromatographic analysis with direct-injection-method was used. A well defined volume with a known concentration of the anaesthetic was added to a vessel with known volume. After equilibration the concentration of the inhalational anaesthetics was determined in the liquid at various temperatures. The solubility coefficient and the temperature coefficient were then calculated from these data. We obtained the following solubility coefficients at -4 degrees C (37 degrees): for halothane 1,07 (0,77), for enflurane 1,14 (0,70), for methoxyflurane 4,92 (3,23) and for isoflurane 1,13 (0,82). In Schindler's liquid culture modified by Karzel the values at 24 degrees (37 degrees) were: for halothane 1,33 (0,92), for enflurane 1,32 (0,87), for methoxyflurane 6,61 (4,48) and for isoflurane 1,19 (0,98). The temperature coefficient for this temperature range were in water (in Schindler's liquid culture) for halothane: -2,31 . 10(-2) K(-1) (-3,15 . 10(-2) K(-1)), for methoxyflurane: -13,0 . 10(-2) K(-1) (-16,48 . 10(-2) K(-1)), for enflurane: -3,38 . 10(-2) K(-1) (-3,46 . 10(-2) K(-1)) and for isoflurane: -2,38 . 10(-2) K(-1) (-1,62 . 10(-2) K(-1)).     

Skip metastases: analysis of their clinical significance and prognosis in the IIIA stage of non-small cell lung cancer.

OBJECTIVE: To determine the clinical significance of skip metastases (pN2/S) in patients with resected pIIIA/N2 NSCLC. The incidence of pN2/S after mediastinal lymph node dissection (MLD) and sampling (MLS) were compared. METHOD: From 1997 to 2000, 580 lung resections for NSCLC performed at our department. The 151 patients (26.5%) at stage IIIA/N2 (pN2+) were grouped according to their skip metastases status. Group A included the ordinary pN2 (pN2/O) cases (71%) and group B the pN2/S (29%). Age, gender, type of resection, right or left lesion, histology, tumor lobe predilection, MLD or MLS pathologic results, the level and the number of node stations involved and survival were analyzed. RESULTS: In 44 patients (29%) pN2/S disease was present. Statistical analysis revealed significant difference between pN2/O and pN2/S for the following: (1) pN2/S was more common for right-sided lesions (P=0.007); (2) Squamous carcinoma was the main type of pN2/S (P=0.007) and (3) pN2/S was more frequently detected after MLD than after MLS (P=0.001). Although pN2/S involved more often upper mediastinal lymph nodes (Nos. 2, 3) and one station level (pN2/S: 41 vs. pN2/O: 19.6%, P=0.228) was not found statistically significant. pN2/O was more common after right upper lobectomy and pN2/S after right lower lobectomy. The 3-year survival was more favorable for the pN2/S group (A: 24 vs. B: 36%, P=0.07). CONCLUSION: (1) MLD was found to be more reliable for pN2/S detection than MLS. (2) The presence of pN2/S proved to be a less aggressive form of lymphatic spread that should be taken into account in the future. (3) Strong correlation between right lower lobe tumors and pN2/S was demonstrated. (4) Different routes of cancer lymphatic spread between pN2/S and pN2/O are suggested.     

2-chloroprocaine antagonism of epidural morphine analgesia

Background: 2-chloroprocaine (2-CP) used for lumbar epidural anesthesia (LEA) reportedly decreases the efficacy of epidural morphine (EM) administered for post-cesarean section (CS) analgesia. The amount of supplemental i.v. morphine self-administered by the patient via the patient-controlled analgesia device (PCA) is used to study the interaction between EM and 2-CP.
Methods: Forty-two patients scheduled for elective CS were randomly divided into 3 equal groups, and received 2-CP, 2-CP+epinephrine (Epi, 5 μg ml^-1) or 2% lidocaine (Lido) with Epi for LEA. All patients received 5 mg EM and i.v. PCA morphine for postoperative pain. Cumulative amount of i.v. morphine used in the first 24 hours as well as the amount of the drug used during each 2-h period were noted. Nonparametric analysis of variance and Chi-squared analysis were used for statistical comparisons.
Results: The mean cumulative 24-h i.v. PCA morphine requirement in the 2-CP, 2-CP+Epi and Lido+Epi groups respectively was 20.5±24, 33.1.5±27 and 4.07±6.3 (mean±SD). The Lido+Epi group used significantly less morphine ( P = 0.01) compared to either of the 2-CP groups with no significant difference between the 2-CP groups. The maximum i.v. PCA morphine use occurred in the first 4 hours following surgery in all three groups.
Conclusion: Analgesic efficacy of EM is decreased when 2-CP is used for LEA compared to when Lido+Epi is used.     

Comparison of different near-infrared spectroscopic cerebral oxygenation indices with central venous and jugular venous oxygenation saturation in children

BACKGROUND: We compared two different near-infrared spectrophotometers: cerebral tissue oxygenation index (TOI) measured by NIRO 200 and regional cerebral oxygenation index (rSO(2)) measured by INVOS 5100 with venous oxygen saturation in the jugular bulb (SjO(2)) and central SvO(2) from the superior caval vein (SVC) during elective cardiac catheterization in children. METHODS: A prospective observational clinical study in 31 children with congenital heart defects in a catheterization laboratory was undertaken. TOI was compared with SjO(2) in the left jugular bulb and with SvO(2). rSO(2) was compared with SjO(2) from the right jugular bulb and SvO(2). Linear regression analysis and Pearson's correlation coefficient were calculated and Bland-Altman analyses were performed. RESULTS: Cerebral TOI and SjO(2) were significantly correlated (r = 0.56, P < 0.0001), as well as TOI and SvO(2) with r = 0.74 (P < 0.0001). Bland-Altman plots showed a mean bias of -4.3% with limits of agreement of 15.7% and -24.3% for TOI and SjO(2) and a mean bias of -4.9% with limits of agreement of 10.3% and -20.1% for TOI and SvO(2). Cerebral rSO(2) and SjO(2) showed a significant correlation (r = 0.83, P < 0.0001) and rSO(2) and SvO(2) showed excellent correlation with r = 0.93 (P < 0.0001). Bland-Altman plots showed a mean bias of -5.2% with limits of agreement of between 8.4% and -18.8% for rSO(2) and SjO(2) and a mean bias of 5.6% with limits of agreement of 13.4% and -2.2% for rSO(2) and SvO(2). CONCLUSIONS: Both near-infrared spectroscopy devices demonstrate a significant correlation with SjO(2) and SvO(2) values; nevertheless both devices demonstrate a substantial bias of the measurements to both SjO(2) and SvO(2).     

Characterization of sulphate transporters in isolated bovine articular chondrocytes.

Uptake of SO(4) (2-) by articular chondrocytes is an essential step in the pathway for sulphation of glycosaminoglycans (GAGs), with mutations in SO(4) (2-) transport proteins resulting in abnormalities of skeletal growth. In the present study, the transporters mediating SO(4) (2-) transport in bovine articular chondrocytes have been characterized. Expression of candidate transporters was determined using RT-PCR, while SO(4) (2-) transport was measured in radioisotope flux experiments. RT-PCR experiments showed that bovine articular chondrocytes express three transporters known to transport SO(4) (2-): AE2 (SLC4a2), DTDST (SLC26a2), and SLC26a11. Other transporters--NaS-1 (SLC13a1), SAT-1 (SLC26a1), DRA (SLC26a3), SLC26a6 (PAT1), SLC26a7, SLC26a8 (Tat-1), and SLC26a9--were, however, not detected. In functional experiments, SO(4) (2-) uptake was temperature-sensitive, inhibited by 60% by DIDS (50 microM) and exhibited saturation kinetics, with a K(m) value of 16 mM. Uptake was also inhibited at alkaline extracellular pH. In further experiments, a K(i) value for DIDS inhibition of SO(4) (2-) efflux of 5 microM was recorded. A DIDS-sensitive component of SO(4) (2-) efflux persisted in solutions lacking Cl(-) ions. These data are interpreted as evidence for the preferential operation of carrier-mediated exchange of SO(4) (2-) for Cl(-), while an alternative SO(4) (2-)-OH(-) exchange mode is also possible.     

A comparative evaluation of transcutaneous and end-tidal measurements of CO2 in thoracic anesthesia

We performed this study to assess the accuracy of transcutaneous CO(2) (PTCCO(2)) monitoring compared with end-tidal CO(2) (PETCO(2)) in thoracic anesthesia. Twenty-six patients undergoing pneumonectomy with thoracotomy for which a long period of one-lung ventilation (OLV) was required were studied. The lungs were mechanically ventilated in the lateral decubitus position. PTCCO(2), PETCO(2), and arterial CO(2) (PaCO(2)) were simultaneously measured during two-lung ventilation (TLV) and during OLV at intervals of 15 min. All patients completed the study protocol. Bland-Altman analysis revealed a bias of -0.4 mm Hg with a precision of +/-2.5 mm Hg during OLV and 1.4 mm Hg with +/-4.3 mm Hg during TLV when PTCCO(2) and PaCO(2) were compared and revealed a bias of -5.8 mm Hg with a precision of +/-4.1 mm Hg during OLV and -7.1 mm Hg with +/-4.6 mm Hg during TLV when PETCO(2) and PaCO(2) were compared. We conclude that PTCCO(2) monitoring is accurate for evaluating CO(2) levels during thoracic anesthesia.     

A pilot study of continuous transtracheal mixed venous oxygen saturation monitoring

In this study, we investigated the feasibility and the accuracy of transtracheal mixed venous oxygen saturation (Svo(2)) monitoring. Ten patients undergoing thoracic surgery were included in this study. A single-use pediatric pulse oximetry sensor was attached to the double-lumen tube between the tracheal and bronchial cuff. After anesthesia was induced, the double-lumen tube was inserted into the trachea and adjusted to the proper position. During surgery, the pulmonary arterial blood was sampled every 3 min for 15 min to measure the Svo(2). The measurements made by the transtracheal pulmonary pulse oximeter (Sto(2)) were recorded at the same time that blood was sampled from the pulmonary artery for Svo(2) measurements. The levels of measurement agreement between the Sto(2) and the Svo(2) were analyzed using the Bland and Altman method. The mean +/- sd (range) oxygen saturation values during the data collecting period were 82.0% +/- 4.9% (72%-91%) for the Sto(2) and 82.2% +/- 5.5% (71%-91%) for the Svo(2), respectively. The linear correlation coefficient of the regression analysis between the Sto(2) and the Svo(2) was 0.934 (P < 0.05). A 95% confidence interval for absolute difference between the Sto(2) and the Svo(2) was 1.58%-2.09%. The mean +/- 2 sd difference between the Sto(2) and the Svo(2) was 0.12% +/- 3.97% on the Bland and Altman graph. We conclude that it is feasible to monitor the pulmonary artery oxygen saturation continuously by a transtracheal pulse oximetry technique and that it can be done so accurately. IMPLICATIONS: Mixed venous oxygen saturation (Svo2) is a measure of the balance between oxygen supply and consumption throughout the whole body. Svo2 can be measured invasively by inserting a pulmonary artery catheter with the associated disadvantages of cost and potential for patient injury. In this study, we investigated the feasibility of noninvasive Svo2 measurement using a transtracheal pulse oximetry technique.     

Effect of 1,25-dihydroxyvitamin D3 on mouse mammary tumor (GR) cells: evidence for receptors, cellular uptake, inhibition of growth and alteration in morphology at physiologic concentrations of hormone

Mammary glands are target tissues for 1,25-dihydroxyvitamin D3 (1,25(OH)2D3). We have examined a mouse mammary tumor cell line (GR) for receptors of 1,25(OH)2D3 and have examined alterations in the growth and morphology of these cells in response to 1,25(OH)2D3. GR cells contain a high affinity (Kd approximately 10(-11)), low-capacity receptor with a high specificity for 1,25(OH)2D3. The 1,25(OH)2D3 receptor in GR cells has a sedimentation coefficient of 3.5 and elutes from DEAE cellulose columns with approximately 0.15 M KCl. These properties of the receptor are similar to those reported for other 1,25(OH)2D3 receptors. 1,25(OH)2D3 is internalized by GR cells in situ and specifically bound 1,25(OH)2D3 is found predominantly, if not entirely, in the nucleus as determined by cell fractionation and autoradiographic techniques. The incubation of GR cells in culture for 7 days with 1,25(OH)2D3 markedly alters cell growth. Cell growth is retarded in a dose-dependent manner; physiologic concentrations (10(-10) M) of 1,25(OH)2D3 retard cell growth by approximately 50%. In addition, GR cells incubated with 10(-9) to 10(-8) M 1,25(OH)2D3 undergo marked morphological changes. The incubation of GR cells with other vitamin D metabolites such as 25-hydroxyvitamin D3 (25(OH)D3) at a concentration of 10(-9) M does not significantly alter cell growth or morphology. The presence of high affinity receptors for 1,25(OH)2D3, the specific internalization of 1,25(OH)2D3 predominantly into the nuclei, and the significant effects of physiological concentrations of 1,25(OH)2D3 on cell growth suggest a direct, specific, nuclear effect of 1,25(OH)2D3 on GR cells. The mouse mammary tumor model might be useful in examining the effect of 1,25(OH)2D3 on tumor formation.     

Rapid deflation of the bronchial cuff of the double-lumen tube after decreasing the concentration of inspired nitrous oxide

Deflationary phenomena of the endotracheal tube cuff may occur after inspired nitrous oxide (N(2)O) concentrations are reduced, but deflationary phenomena of the double-lumen tube (DLT) cuff have not been investigated. In this study, tracheal and bronchial cuffs of left-sided Mallinckrodt (Athlone, Ireland) DLTs were inflated with air, 40% N(2)O, or 67% N(2)O (Air, N40, or N67 groups, respectively) in 24 patients undergoing thoracic surgery; 40 min later, O(2) was substituted for N(2)O in some of the patients in the N40 group (N40-c group). Intracuff gas volumes, N(2)O concentrations, and cuff compliance were also measured. Both tracheal and bronchial cuff pressures significantly increased in the Air group but decreased in the N67 group. Neither pressure significantly changed in the N40 group, but both decreased in the N40-c group after terminating N(2)O anesthesia; the time required for bronchial cuff pressures to decrease by half (12.0 +/- 5.5 min) was less than that for tracheal cuff pressures (31.2 +/- 11.0 min, P < 0.01). The volume change in the N40-c group was not significantly different between the tracheal and bronchial cuffs, but tracheal cuff compliance was significantly higher than bronchial compliance. Therefore, filling DLT cuffs with 40% N(2)O stabilizes cuff pressure during anesthesia with 67% N(2)O, but bronchial cuffs deflate more quickly than tracheal cuffs after cessation of N(2)O administration through smaller compliance. IMPLICATIONS: We demonstrated that after cessation of nitrous oxide (N(2)O) administration, bronchial N(2)O-filled cuffs of the double-lumen tube deflate more rapidly than tracheal cuffs. To avoid insufficient separation of the lungs by the bronchial cuff, a frequent check of the cuff pressure is recommended after the inspired N(2)O concentration is decreased.