中医院校研究生药物研究的实验设计

实验设计是整个科学研究工作的基石,实验设计不合理或不正确,轻则造成人力物力的浪费,重则导致整个科研的失败.本文从多年的中医学院科研教学经验中,总结出研究生进行药物研究时实验设计的几点注意事项,为相关领域提供参考.     

Inhalation experiments with mixtures of hydrocarbons Experimental design,statistics and interpretation of kinetics and possible interactions

 The paper describes experimental and statistical methods for toxicokinetic evaluation of mixtures in inhalation experiments. Synthetic mixtures of three C9 n-paraffinic, naphthenic and aromatic hydrocarbons (n-nonane, trimethylcyclohexane and trimethylbenzene, respectively) were studied in the rat after inhalation for 12 h. The hydrocarbons were mixed according to principles for statistical experimental design using mixture design at four vapour levels (75, 150, 300 and 450 ppm) to support an empirical model with linear, interaction and quadratic terms (Taylor polynome). Immediately after exposure, concentrations of hydrocarbons were measured by head space gas chromatography in blood, brain, liver, kidneys and perirenal fat. Multivariate data analysis and modelling were performed with PLS (projections to latent structures). The best models were obtained after removing all interaction terms, suggesting that there were no interactions between the hydrocarbons with respect to absorption and distribution. Uptake of paraffins and particularly aromatics is best described by quadratic models, whereas the uptake of the naphthenic hydrocarbons is nearly linear. All models are good, with high correlation (r ²) and prediction properties (Q ²), the latter after cross validation. The concentrations of aromates in blood were high compared to the other hydrocarbons. At concentrations below 250 ppm, the naphthene reached higher concentrations in the brain compared to the paraffin and the aromate. Statistical experimental design, multivariate data analysis and modelling have proved useful for the evaluation of synthetic mixtures. The principles may also be used in the design of liquid mixtures, which may be evaporated partially or completely. Received: 26 June 1995/Accepted: 17 October 1995     

用SAS软件实现交叉设计定量资料的统计分析

交叉设计是药学科研中研究两种或多种药物疗效时使用的一种实验设计方法,在临床药物疗效研究方面具有十分广泛的适用性。本文主要从交叉设计的概念、统计分析方法的合理选用、如何使用SAS程序进行分析,以及如何解释结果几个方面作一概述。     

用SAS软件实现析因设计定量资料的统计分析

析因设计是医药学科研中经常用到的一种实验设计方法,在药物疗效研究方面具有十分广泛的适用性,用于考察两个及以上实验因素及其交互作用对实验结果的影响。本文主要从析因设计的概念、析因设计统计分析方法的合理选用、如何使用SAS程序进行分析,以及如何解释结果几个方面作一概述。     

Statistical optimization applied to the spectrophotometric study of a tolmetin—copper(II) complex

Tolmetin sodium has been investigated and determined from dosage forms as its Cu(II) complex and method optimized by statistical optimization. The assay was developed using two mathematical statistical models: factorial design and response-surface mapping. The decision to apply experimental design techniques to the development of the method was made after a series of screening experiments revealed that the complex formation and extraction are maximized as a function of supporting electrolyte concentration, concentration of Cu(II) acetate and pH of the reaction mixture. One set of two-level three variable factorial experiments was carried out in order to evaluate the main effect, as well as the interaction among factors. The final step was to optimize the values of variables using response surface design. The best set of conditions was selected for further investigation.     

Modeling Methods for Mixture-of-Mixtures Experiments Applied to a Tablet Formulation Problem

During the past few years, statistical methods for the experimental design, modeling, and optimization of mixture experiments have been widely applied to drug formulation problems. Different methods are required for mixture-of-mixtures (MoM) experiments in which a formulation is a mixture of two or more “major” components, each of which is a mixture of one or more “minor” components. Two types of MoM experiments are briefly described. A tablet formulation optimization example from a 1997 article in this journal is used to illustrate one type of MoM experiment and corresponding empirical modeling methods. Literature references that discuss other methods for MoM experiments are also provided.     

均匀设计方法及其应用

利用统计学方法处理科学试验数据是当今在科学研究中重要的一个环节 ,但应当注意到在试验过程中试验设计是否合理直接影响到试验的结果和统计分析。另一方面 ,科学的试验设计可以大量的节省试验次数 ,达到试验的最佳效果。通过对均匀设计与正交设计的比较 ,说明了均匀设计可用较少的试验次数达到较好的试验效果 ,同时指出了其在应用中应注意的问题。     

The use of the Box-Behnken experimental design in the optimisation and robustness testing of a capillary electrophoresis method for the analysis of ethambutol hydrochloride in a pharmaceutical formulation

Box–Behnken experimental designs do not appear to be extensively used in optimisation of analytical methods using capillary electrophoresis (CE). This paper describes the use of the Box–Behnken experimental design to optimise the factors affecting the separation of ethambutol hydrochloride (EB), its impurity 2-amino-1-butanol and the internal standard (phenylephrine hydrochloride) in a CE method for a pharmaceutical tablet assay. The three factors studied simultaneously were: buffer pH, buffer concentration and applied electric field, each at three levels. The method was optimised with respect to three responses: resolution between peaks, theoretical plate count and the migration time of the EB peak. A statistical programme, which applies a multiple response optimisation algorithm, was used to calculate and optimise the three responses simultaneously. The optimum conditions were established to be 58.0 mM sodium borate buffer at pH 9.50 and an applied electric field of 412 V/cm. The robustness of the method was also determined and confirmed using a second Box–Behnken design, as part of the validation exercise. System suitability values for the method were derived from the regression surface analysis. The CE method for a pharmaceutical tablet formulation was further validated according to current regulatory requirements, with respect to linearity and range, precision, specificity, accuracy and limit of quantitation. The optimised method gives a fast and efficient separation under 4 min, with complete resolution between the three peaks, and represents an improvement over the existing USP method. It can be concluded that the Box–Behnken experimental design provides a suitable means of optimising and testing the robustness of a CE pharmaceutical method.     

基于创新能力提升的药学开放性实验教学平台研究

药学是实践性很强的学科,创新能力培养成为高等学校的重要任务之一,开放性创新实验教学则是以学生为主导作用、学生根据所学的理论知识自己设计实验、实施实验、整理总结实验结果和撰写研究报告／论文的一种实验模式,是激发学生的科研兴趣、培养学生的科研素质、提高学生的动手力和创新能力的富有个性的实验教学模式。因此,构建基于创新能力提升的药学开放性实验教学平台具有十分重要的意义。     

Experimental design and capillary electrophoresis for simultaneous analysis of arbutin, kojic acid and hydroquinone in cosmetics

A statistical experimental design was used to optimize one micellar electrokinetic capillary electrophoresis (MEKC) for simultaneous analysis of arbutin (AR), kojic acid (KA) and hydroquinone (HQ). Untreated fused-silica capillaries were operated using a phosphate buffer (20mM, pH 6.5) under 20 kV and detection at 200 nm. Quantitative linear ranges were 20-200 microg/ml for AR, 20-100 microg/ml for KA and 8-80 microg/ml for HQ with correlation coefficients >or=0.9994. R.S.D. and R.E. were less than 3.0% for the intra-day and inter-day analysis, and all recoveries were greater than 99%. Our method was applied to assay commercial cosmetics. The results were within the labeled amount of 99.6-102.5%.     

Sequential Experimentation in Product Development. I

Investigations proceed as an iterative and recursive learning process, involving a sequence of experiments. Evaluating the results from a prior set of experiments allows the investigator to design and predict the next experimental cycle better. The investigator looks at what has gone before with a view to improve and extend the inquiry. This communication reiterates the value of sequential experimentation in product development. A fluid-bed granulation study is used to illustrate the sequential approach. Acceptable solutions were achieved using technical knowledge, a sequence of factorial designs, and graphical data analysis.     

An investigation of the influence of process and formulation variables on mechanical properties of high shear granules using design of experiment

Being able to predict the properties of granules from the knowledge of the process and formulation variables is what most industries are striving for. This research uses experimental design to investigate the effect of process variables and formulation variables on mechanical properties of pharmaceutical granules manufactured from a classical blend of lactose and starch using hydroxypropyl cellulose (HPC) as the binder. The process parameters investigated were granulation time and impeller speed whilst the formulation variables were starch-to-lactose ratio and HPC concentration. The granule properties investigated include granule packing coefficient and granule strength. The effect of some components of the formulation on mechanical properties would also depend on the process variables used in granulation process. This implies that by subjecting the same formulation to different process conditions results in products with different properties.     

Optimization and validation of a CZE method for rufloxacin hydrochloride determination in coated tablets

A simple and rapid capillary electrophoresis method with UV detection was developed and validated for the determination of rufloxacin hydrochloride in coated tablets. An experimental design strategy (Doehlert design and desirability function) allowed the analytical parameters to be simultaneously optimized in order to determine rufloxacin hydrochloride with high peak area/migration time ratio, good efficiency and short analysis time. Optimized analyses were run using boric acid 0.10 M adjusted to pH 8.8 as BGE and setting voltage and temperature at 18 kV and 27 °C, respectively. Pefloxacin mesylate was used as internal standard and run time was about three minutes. The method was validated for the drug substance and the drug product according to the ICH3 guidelines. Robustness was tested by experimental design using an eight-run Plackett–Burman matrix.     

用SAS软件实现随机区组设计定量资料的统计分析

随机区组设计是药学科研中控制重要的非试验因素时用到的一种试验设计方法,在临床药物疗效研究方面具有十分广泛的适用性.本文主要对随机区组设计的概念,统计分析方法的合理选用,如何使用SAS程序进行分析,以及如何解释结果几个方面作一概述.     

The effect of experimental design on the modeling of a tablet coating formulation using artificial neural networks

The aim of this study was to investigate the effect of experimental design strategy on the modeling of a film coating formulation by artificial neural networks (ANNs). Box-Behnken, central composite and pseudo-random designs of 102, 90 and 100 simulated records, respectively were used to train a multilayer perceptron (MLP) ANN comprising six input and two output nodes separated by a single hidden layer of five nodes. Network over-training was limited by using a test set of 40 pseudo-randomly distributed records. The models were validated using a set of 60 pseudo-randomly distributed records. Crack velocity was highly curved with respect to pigment particle size and size distribution. Similarly, film opacity was highly curved in response to pigment concentration and film thickness. The Box-Behnken and central composite designs generated models that were unable to predict crack velocity and showed extensive bias in prediction of film opacity. The pseudo-random design was unable to predict crack velocity of the test data set but yielded acceptable predictions for the validation set. Film opacity was well predicted by the pseudo-random design model. The poor predictive ability of the Box-Behnken and central composite models was attributed to poor interpolation of the high curvature of the response surfaces. In contrast, the pseudo-random design mapped the interior of the design space allowing improved interpolation and predictive ability. It is concluded that Box-Behnken and central composite experimental designs are inappropriate for ANN modeling of highly curved responses and that extensive internal mapping of the design space is essential to generate predictive ANN models.     

Mixture experiment methods in the development and optimization of microemulsion formulations

Microemulsion formulations represent an interesting delivery vehicle for lipophilic drugs, allowing for improving their solubility and dissolution properties. This work developed effective microemulsion formulations using glyburide (a very poorly-water-soluble hypoglycaemic agent) as a model drug. First, the area of stable microemulsion (ME) formations was identified using a new approach based on mixture experiment methods. A 13-run mixture design was carried out in an experimental region defined by constraints on three components: aqueous, oil and surfactant/cosurfactant. The transmittance percentage (at 550 nm) of ME formulations (indicative of their transparency and thus of their stability) was chosen as the response variable. The results obtained using the mixture experiment approach corresponded well with those obtained using the traditional approach based on pseudo-ternary phase diagrams. However, the mixture experiment approach required far less experimental effort than the traditional approach. A subsequent 13-run mixture experiment, in the region of stable MEs, was then performed to identify the optimal formulation (i.e., having the best glyburide dissolution properties). Percent drug dissolved and dissolution efficiency were selected as the responses to be maximized. The ME formulation optimized via the mixture experiment approach consisted of 78% surfactant/cosurfacant (a mixture of Tween 20 and Transcutol, 1:1, v/v), 5% oil (Labrafac Hydro) and 17% aqueous phase (water). The stable region of MEs was identified using mixture experiment methods for the first time.     

用SAS软件实现多因素重复测量设计定量资料的统计分析

重复测量是药学科研临床实验中经常用到的一种实验设计方法,在临床药物疗效研究方面具有十分广泛的适用性.本文主要从重复测量的概念、重复测量统计分析方法的合理选用、如何使用SAS程序进行分析,以及如何解释结果几个方面作一概述.     

用SAS软件实现横断面设计定性资料的统计分析

横断面设计是药学科研中经常用到的一种实验设计方法,在临床药物疗效研究方面具有十分广泛的适用性。本文主要从横断面设计的概念、分析方法的合理选用、如何使用SAS程序进行分析,以及如何解释结果几个方面作一概述。     

Uncertainty of measurement and error in stability studies

It is required that shelf life be determined based on the lower limit of the confidence interval of the estimate from the stability tests. Simulations indicate that a 1-year prediction of shelf life will have approximately 1 month of error. However, this is product specific and is related to the uncertainty of measurement and experimental design. Factors associated with product and experimental design, such as degradation rate, number of time points, implementing a full versus a reduced design, etc., can significantly affect the error of shelf life. Uncertainty in measurement is positively correlated to the amount of error through the manufacturing lot-to-lot variability, precision of the analytical method and calibrator. Experimental design can control random variability and actually can reduce error by increasing number of lots and replicates in stability tests. The decision on the number of lots and replicates will be a balancing act between the uncertainty of the measurement, design and other practical considerations.     

试验设计的三要素

试验研究中不可缺少的三项内容分别是试验因素、受试对象和试验效应,通常将他们称为试验设计的三要素。如何正确把握三要素是试验设计的关键问题之一。