线粒体和亚线粒体体外代谢及漆酶影响的微量热研究

首次将微量热用于亚细胞层次的研究，测定了鲤鱼肝脏线粒体和亚线粒体代谢的热谱及其漆树漆酶的影响，并用ＭＴＤ方程和演化计算技术对它们的代谢热动力学参数进行了全局优化。     

量热法研究线粒体代谢的热力学和动力学行为

线粒体是细胞中极为重要的细胞器，是产生细胞所必需的“富能”物质ATP的重要场所，为细胞活动提供所需化学能．在生命体能量代谢过程中除一部分能量用于合成ATP外，其余则以热的形式释出．用精密热量计测出线粒体代谢过程中的热量输出对了解线粒体的功能和代谢机制具有十分重要的意义．用微量热法研究线粒体体外代谢已有一些报导［‘，’］．本工作用精密热量计和差式扫描量热仪侧定了水稻线粒体体外代谢热谱和DSC曲线，计算了水稻线粒体活性增长速率常数，比较了不同保藏时间的水稻线粒体体外代谢的差异，并初步探讨了水稻线粒体在变…     

不同条件下离体大鼠肝脏线粒体能量代谢微量热分析

为了探究线粒体的能量代谢过程,本文以离体大鼠肝脏线粒体为模型,利用多通道、高灵敏度的热活性检测仪TAM Ⅲ,实时监测了不同线粒体浓度、不同底物、不同缓冲液、几种呼吸抑制剂以及Ca²⁺和线粒体渗透转换孔抑制剂CsA存在时线粒体的能量代谢,获得了完整的热功率―时间曲线,并通过计算得到了线粒体能量代谢的热动力学参数。通过分析发现：（1）线粒体浓度越大,代谢越快;（2）直接底物琥珀酸钠使线粒体代谢更快;（3）高浓度Ca²⁺能够刺激线粒体快速产热,且在长期代谢进程中,线粒体渗透转换孔抑制剂CsA并不能改变Ca²⁺造成的影响;（4）不同缓冲液对线粒体代谢的影响基于其组分的不同,缓冲液中含有呼吸底物;（5）呼吸抑制剂都能抑制线粒体的能量代谢,尤其是复合物IV的抑制剂NaN₃,高浓度下使代谢停止。     

等温微量热法在生命科学研究中的应用

简要介绍了等温微量热法的原理、典型的仪器及其在生命科学研究中应用所具有的特点。通过它可获得完整的细胞代谢过程产热曲线及其热动力学方程；可以研究细胞器的代谢规律；可以获取生物大分子与小分子相互作用的热动力学特征。     

线粒体体外代谢热动力学研究

线粒体是细胞重要的细胞器之一，有细胞的“能源工厂”之称．因为线粒体内有许多酶，是特殊的酶催化氧化反应的场所山；所有动、植物细胞的线粒体都能通过各种营养物的氧化而产生“富能”物质ATP．采用一定的技术可将线粒体从细胞中分离出来，分离出来的线粒体中的酶系统还有一定的活性，而且线粒体内也有一定的营养物质，这样酶系统就能利用这些营养物进行代谢，从而释放出一定的能量．我们用微量热法对两种鱼肝脏线粒体进行了测量，发现线粒体代谢过程分四个阶段：停滞期、活性恢复期、稳定期、活性衰减期．在活性恢复期和活性衰减期，…     

热动力学特征对比参量法及其应用研究

基于简单级数反应的积分和微分热动力学方程，建立了热动力学特征对比参量 法的数学模型，提出了一种由特征时间参量计算特征对比参量的方法，通过几种不 同级数模型反应的热动力学研究证明了方法的正确性，并利用该法研究了过氧化氢 在磷酸盐缓冲液中氧化对苯醌反应的动力学特征，实验结果表明该反应动力学方程 可以表示为：dC(醌)/dt=kC(醌)C(H_2O_2)~0.5C(H~+)~(-0.5)     

模拟热谱曲线法（Ⅰ）——简单级数化学反应

提出模拟各种简单级数化学反应的普适热谱曲线方程△＝ａｔｅ＾－ｋβｔ，建立了一种新的热动力学研究法－－模拟热谱曲线法，导出了简单级数化学反应的动力学参数Ｋｎ和速率常数ｋｎ的计算式。实验说明该方法对研究慢反应及较快的反应均适用，还能用于求热动力学体系的冷却常数。     

酶促反应抑制动力学的微量热法研究——氟离子对漆树漆酶催化…

提出一个适合各类可逆性抑制的热动力学方程，并根据各量之间的关系提出了可逆竞争性抑制、非竞争抑制、反竞争性抑制的热动力学判据以及计算表观米氏常数Ｋｍ，ａｐｐ和抑制常数Ｋｉ等酶促反应生化常数的热动力学公式，并实验应用于ＮａＦ对漆树漆酶催化氧化邻苯二胺抑制向量热法温室，实验初步得出此抑制为非竞争性抑制，求出其Ｋｍ，ａｐｐ，Ｋｉ值。     

平行反应的热动力学研究法

根据热动力学基本理论, 推导了平行准一级反应和平行准一二级反应的热动力学方程, 建立了热谱解析平行反应动力学参数的热动力学研究法, 并利用该法研究了两个模拟平行反应体系的热动力学, 实验结果验证了方法的正确性.     

细菌生长的热动力学研究进展

本文简单介绍了绝热式和热导式量热计及理论模型,细菌生长的动力学模型,着重介绍了近五年中热动力学在研究细菌生长代谢过程中的应用;通过测定不同条件下细菌生长代谢过程的产热曲线,求得细菌生长代谢的热力学和动力学参数,研究细菌的生长条件和药物的抑制作用。     

Replica state exchange metadynamics for improving the convergence of free energy estimates

Metadynamics (MTD) is a powerful enhanced sampling method for systems with rugged energy landscapes. It constructs a bias potential in a predefined collective variable (CV) space to overcome barriers between metastable states. In bias‐exchange MTD (BE‐MTD), multiple replicas approximate the CV space by exchanging bias potentials (replica conditions) with the Metropolis–Hastings (MH) algorithm. We demonstrate that the replica‐exchange rates and the convergence of free energy estimates of BE‐MTD are improved by introducing the infinite swapping (IS) or the Suwa‐Todo (ST) algorithms. Conceptually, IS and ST perform transitions in a replica state space rather than exchanges in a replica condition space. To emphasize this, the proposed scheme is called the replica state exchange MTD (RSE‐MTD). Benchmarks were performed with alanine polypeptides in vacuum and water. For the systems tested in this work, there is no significant performance difference between IS and ST. © 2015 Wiley Periodicals, Inc.     

Development of Simple Electrochemical Sensor for Selective Determination of Methadone in Biological Samples Using Multi‐walled Carbon Nanotubes Modified Pencil Graphite Electrode

The electrochemical sensor was developed for determination of methadone (MTD) using multi‐walled carbon nanotubes (MWCNT) modified pencil graphite electrode (MWCNT‐PGE). It was found that the oxidation peak current of MTD at the MWCNT‐PGE was greatly improved compared with that of the bare‐PGE. At the MWCNT‐PGE, well‐defined anodic peak of MTD was observed at about 0.7 V (in pH 7 solution). The influence of several parameters on the determination of MTD was investigated. At optimum experimental conditions, differential pulse voltammetry (DPV) was used for determination of MTD, which exhibited a linear calibration graph of Ip versus MTD concentration in the range of 0.1–15 µM with a correlation coefficient of 0.9992. The calculated detection limit for S/N = 3 was 87 nM. It has been shown that the peaks obtained for oxidation of ascorbic acid (AA), uric acid (UA) and MTD in their mixture could be well resolved by differential pulse voltammetry, permitting us to develop a sensitive and selective electrochemical sensor for determination of MTD in the presence of AA and UA. Finally, MWCNT‐PGE was used for determination of MTD in biological samples, such as human serum and urine, using the standard addition procedure and the results were quite promising.     

Steric and electrostatic effects in dye-cellulose interactions by the MTD and CoMFA approaches

This paper presents the application of the MTD (minimal steric difference) analysis and CoMFA (comparative molecular field analysis) to series of anthraquinone vat, mono and disazo and disperses dyes with known affinities for cellulose fiber. A comparison of the results demonstrates that these methods usually agree with the prediction of structural features favorable for dyeing. A series of n = 49 anthraquinone vat dyes was studied by MTD with r2 between 0.903 and 0.941 and r2CV values in the range of 0.827-0.878. For CoMFA, r2 = 0.992, r2CV = 0.841 were obtained; the CoMFA field is in rather good agreement with vertex attributions, by MTD for attractive and repulsive vertices. Anionic disazo dyes were studied by the CoMFA method (n = 21, r2 = 0.999, r2CV = 0.703). Monoazo dyes (several series) were studied by CoMFA and MTD. The effect of lipophilicity on dye fiber affinity was, also, studied for these dyes. Disperse dye adsorption was analyzed by MTD and CoMFA (n = 27, r2 = 0.925, r2CV = 0.776). Conclusions refer to the effect of structural features of dye molecules upon adsorption on cellulose fibers.     

Steric and electrostatic effects in dye-cellulose interactions by the MTD and CoMFA approaches

This paper presents the application of the MTD (minimal steric difference) analysis and CoMFA (comparative molecular field analysis) to series of anthraquinone vat, mono and disazo and disperses dyes with known affinities for cellulose fiber. A comparison of the results demonstrates that these methods usually agree with the prediction of structural features favorable for dyeing. A series of n =49 anthraquinone vat dyes was studied by MTD with r 2 between 0.903 and 0.941 and r CV 2 values in the range of 0.827-0.878. For CoMFA, r 2 =0.992, r CV 2 =0.841 were obtained; the CoMFA field is in rather good agreement with vertex attributions, by MTD for attractive and repulsive vertices. Anionic disazo dyes were studied by the CoMFA method ( n =21, r 2 =0.999, r CV 2 =0.703). Monoazo dyes (several series) were studied by CoMFA and MTD. The effect of lipophilicity on dye fiber affinity was, also, studied for these dyes. Disperse dye adsorption was analyzed by MTD and CoMFA ( n =27, r 2 =0.925, r CV 2 =0.776). Conclusions refer to the effect of structural features of dye molecules upon adsorption on cellulose fibers.     

Methadone concentrations in blood, plasma, and oral fluid determined by isotope-dilution gas chromatography–mass spectrometry

Methadone (MTD) is widely used for detoxification of heroin addicts and also in pain management programs. Information about the distribution of methadone between blood, plasma, and alternative specimens, such as oral fluid (OF), is needed in clinical, forensic, and traffic medicine when analytical results are interpreted. We determined MTD and its metabolite 2-ethylidene-1,5-dimethyl-3,3-diphenylpyrrolidine (EDDP) in blood, plasma, blood cells, and OF by gas chromatography–mass spectrometry (GC-MS) after adding deuterium-labeled internal standards. The analytical limits of quantitation for MTD and EDDP by this method were 20 and 3 ng/mL, respectively. The amounts of MTD and EDDP were higher in plasma (80.4 % and 76.5 %) compared with blood cells (19.6 % and 23.5 %) and we found that repeated washing of blood cells with phosphate–buffered saline increased the amounts in plasma (93.6 % and 88.6 %). Mean plasma/blood concentration ratios of MTD and EDDP in spiked samples (N?=?5) were 1.27 and 1.21, respectively. In clinical samples from patients (N?=?46), the concentrations of MTD in plasma and whole blood were highly correlated (r?=?0.92, p?<?0.001) and mean (median) plasma/blood distribution ratios were 1.43 (1.41). The correlations between MTD in OF and plasma (r?=?0.46) and OF and blood (r?=?0.52) were also statistically significant (p?<?0.001) and the mean OF/plasma and OF/blood distribution ratios were 0.55 and 0.77, respectively. The MTD concentration in OF decreased as salivary pH increased (more basic). These results will prove useful in clinical and forensic medicine when MTD concentrations in alternative specimens are compared and contrasted.     

Sampling free energy surfaces as slices by combining umbrella sampling and metadynamics

Metadynamics (MTD) is a very powerful technique to sample high‐dimensional free energy landscapes, and due to its self‐guiding property, the method has been successful in studying complex reactions and conformational changes. MTD sampling is based on filling the free energy basins by biasing potentials and thus for cases with flat, broad, and unbound free energy wells, the computational time to sample them becomes very large. To alleviate this problem, we combine the standard Umbrella Sampling (US) technique with MTD to sample orthogonal collective variables (CVs) in a simultaneous way. Within this scheme, we construct the equilibrium distribution of CVs from biased distributions obtained from independent MTD simulations with umbrella potentials. Reweighting is carried out by a procedure that combines US reweighting and Tiwary–Parrinello MTD reweighting within the Weighted Histogram Analysis Method (WHAM). The approach is ideal for a controlled sampling of a CV in a MTD simulation, making it computationally efficient in sampling flat, broad, and unbound free energy surfaces. This technique also allows for a distributed sampling of a high‐dimensional free energy surface, further increasing the computational efficiency in sampling. We demonstrate the application of this technique in sampling high‐dimensional surface for various chemical reactions using ab initio and QM/MM hybrid molecular dynamics simulations. Further, to carry out MTD bias reweighting for computing forward reaction barriers in ab initio or QM/MM simulations, we propose a computationally affordable approach that does not require recrossing trajectories. © 2016 Wiley Periodicals, Inc.