Validation study

Validation can be defined as the act of proving that any procedure, process, equipment, material, activity or system leads to the expected results . Routine and adequate validation studies form a 

All validation procedures must be carefully designed and fully documented in written format (Box 7.2). The results of all validation studies undertaken must also be documented, and retained in the plant files. As part of their routine inspection of manufacturing facilities, regulatory personnel will usually inspect a sample of these records, to ensure conformance to GMP. 

Validation studies encompass all aspects of (bio)pharmaceutical manufacture. All new items of equipment must be validated before being routinely used. Initial validation studies should be 

Validation studies a glossary of some important terms 

Validation master plan Document that serves as an overall guide for a facility s validation programme. It identifies all items/procedures, etc., that must be subjected to validation studies, describes the nature of testing in each instance and defines the responsibilities of those engaged in validation activities 

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C. C. Andeison and E. C. Gundeison, Methods Validation Study of High Peformance Uquid Chromatographic Technique for Determining the MPDA and... 

Validation studies in manufacturing businesses and discussion with experts led to the view that knowledge used to define p, gp, tp and. Vp, could be structured such that may be formulated as ... 

A validation study of the technique showed that it was capable of predicting a high proportion (98%) of errors with serious consequences that actually occurred in an equipment calibration task over a 5-year period (Murgatroyd and Tait, 1987). 

It provides a standardized procedure to ensure consistency among analysts. This was tested by carrying out two independent evaluations of the same task. Of the 60 errors identified in the above validation study, 70% were common to both analysts. Of the remainder, 11 differences were due to differences in knowledge of the equipment by the two analysts and 5 were due to different interpretations of the procedures. 

Cenic A, Nabavi DG, Craen RA, Gelb AW, Lee TY. Dynamic CT measurement of cerebral blood flow a validation study. Am J Neuroradiol 1999 20 63-73. 

Wintermark M, Thiran JP, Maeder P, Schnyder P, Meuli R. Simultaneous measurement of regional cerebral blood flow by perfusion CT and stable xenon CT a validation study. Am J Neuroradiol 2001 22 905-914 [see comment]. 

Baker EL, Letz R, Fidler AT, et al. 1985. A computer-based neurobehavioral evaluation system for occupational and environmental epidemiology Methodology and validation studies. Neurobehav Toxicol Teratol 7 369-377. 

Tam, K. Y., Takacs-Novak, K. Multiwavelength spectroscopic determination of add dissodation constants a validation study. Anal. Chim. Acta 2001, 434,157-167. 

Interest in promoting the use of reference materials in method validation studies led in March 1993 to the formation of a Technical Division on Reference Materials, TDRM (Heavner 1995). The stated purpose of the TDRM is to improve the quality of... 

Until 1991, manufacturers seeking authorizations for pesticides had to fulfil country-specific requirements of validation of enforcement methods. The term enforcement method means analytical methods which are developed for post-registration control and monitoring purposes. The harmonization of these requirements was initiated with the European Economic Community (EEC) Council Directive 91/414/EEC and temporarily finalized with the Guidance Document on Residue Analytical Methods SANCO/825/00 rev. 6, dated 20 June 2000 [Santd et Protection des Consommateurs (SANCO)]. The evaluation of validation studies by the competent authority is conducted by comparison of these European Union (EU) requirements with the study results and most often without any practical experience of the method. Some details of this evaluation are discussed below. 

Consequently, the proof of calibration should never be limited to the presentation of a calibration graph and confirmed by the calculation of the correlation coefficient. When raw calibration data are not presented in such a situation, most often a validation study cannot be evaluated. Once again it should be noted that nonlinearity is not a problem. It is not necessary to work within the linear range only. Any other calibration function can be accepted if it is a continuous function. 

Occasionally the complete sample set of an individual commodity was not analyzed within a validation study. This is not a problem if the same study provides data on additional commodities belonging to the same matrix group. Consequently, the missing data, e.g., a second concentration level, are replaced, provided that control sample results are presented for all crops. 

Better guidance for applicants seems necessary, with the objective that information about extraction efficiency is routinely considered in method validation studies. 

Finally, to avoid the parallel use of similar but not identical method validation studies to fulfil the registration requirements, e.g., of the EU, US Environmental Protection Agency (EPA) or Japanese authorities, an adaptation of different data requirements for residue analytical methods for post-registration control and monitoring purposes would help to save resources. 

For multi-analyte and/or multi-matrix methods, it is not possible to validate a method for all combinations of analyte, concentration and type of sample matrix that may be encountered in subsequent use of the method. On the other hand, the standards EN1528 andEN 12393 consist of a range of old multi-residue methods. The working principles of these methods are accepted not only in Europe, but all over the world. Most often these methods are based on extractions with acetone, acetonitrile, ethyl acetate or n-hexane. Subsequent cleanup steps are based on solvent partition steps and size exclusion or adsorption chromatography on Florisil, silica gel or alumina. Each solvent and each cleanup step has been successfully applied to hundreds of pesticides and tested in countless method validation studies. The selectivity and sensitivity of GC combined with electron capture, nitrogen-phosphorus, flame photometric or mass spectrometric detectors for a large number of pesticides are acceptable. 

The sensitivity achieved (LOD) is not normally presented. It is recognized that different laboratories determine dissimilar values for this parameter and even within a laboratory the repeatability of the LOD is low. Most often, the lowest validated concentration gives an impression about the lowest levels that can be analyzed generally with acceptable results. A measure of selectivity is the intensity of blank results. This intensity is discussed by the participants of inter-laboratory validation studies. However, results are not reported and limits are not defined by CEN TC 275. The results of method validations of the several multi-residue/multi-matrix methods are not reported in the same way, but newer methods with limited scope generate analogous tables with validation results (as an example, see Table 7). 

Each individual method collection comprises a large number of methods, which often have different validation statuses. For instance, the most important Swedish multi-residue method (based on ethyl acetate extraction, GPC and GC) is validated for many pesticides by four laboratories, but other methods are presented with singlelaboratory validation data. Some methods in the Dutch and German manuals were tested in inter-laboratory method validation studies, but others by an independent laboratory or in a single laboratory only. 

If analytical methods are validated in inter-laboratory validation studies, documentation should follow the requirements of the harmonized protocol of lUPAC. " However, multi-matrix/multi-residue methods are applicable to hundreds of pesticides in dozens of commodities and have to be validated at several concentration levels. Any complete documentation of validation results is impossible in that case. Some performance characteristics, e.g., the specificity of analyte detection, an appropriate calibration range and sufficient detection sensitivity, are prerequisites for the determination of acceptable trueness and precision and their publication is less important. The LOD and LOQ depend on special instmmentation, analysts involved, time, batches of chemicals, etc., and cannot easily be reproduced. Therefore, these characteristics are less important. A practical, frequently applied alternative is the publication only of trueness (most often in terms of recovery) and precision for each analyte at each level. No consensus seems to exist as to whether these analyte-parameter sets should be documented, e.g., separately for each commodity or accumulated for all experiments done with the same analyte. In the latter case, the applicability of methods with regard to commodities can be documented in separate tables without performance characteristics. 

The integration of analytical methods in European standards requires their acceptance by several national experts within special working groups and in a final weighted vote of National Standards Bodies. Therefore, there needs to be very high confidence in the performance of methods. Consequently, methods should be tested in inter-laboratory method validation studies, with the exception of those multiresidue methods which are widely used throughout Europe. In the case of CEN methods there is no doubt about residue definition but detailed requirements about the number of matrices and concentration levels in validation experiments do not exist. Eor this reason it may be that CEN methods are validated for important crops only. 

The magnitude of these errors can be analyzed in single laboratories (run bias and repeatability error), in inter-laboratory validation studies (laboratory bias) and in proficiency tests (method bias). Expressed in standard deviations relative to that of... 

Solid-phase microextraction (SPME) consists of dipping a fiber into an aqueous sample to adsorb the analytes followed by thermal desorption into the carrier stream for GC, or, if the analytes are thermally labile, they can be desorbed into the mobile phase for LC. Examples of commercially available fibers include 100-qm PDMS, 65-qm Carbowax-divinylbenzene (CW-DVB), 75-qm Carboxen-polydimethylsiloxane (CX-PDMS), and 85-qm polyacrylate, the last being more suitable for the determination of triazines. The LCDs can be as low as 0.1 qgL Since the quantity of analyte adsorbed on the fiber is based on equilibrium rather than extraction, procedural recovery cannot be assessed on the basis of percentage extraction. The robustness and sensitivity of the technique were demonstrated in an inter-laboratory validation study for several parent triazines and DEA and DIA. A 65-qm CW-DVB fiber was employed for analyte adsorption followed by desorption into the injection port (split/splitless) of a gas chromatograph. The sample was adjusted to neutral pH, and sodium chloride was added to obtain a concentration of 0.3 g During continuous... 

Enforcement method to undergo independent laboratory validation study... 

Lastly, a laboratory not involved in the development process must validate the method. The independent laboratory validation study, or ruggedness trial, ensures that analysts unfamiliar with the method can successfully perform the method. The method developer should, therefore, strive to make all procedures as straightforward as possible to aid reproducibility of the method. 

Multi-residue Method S19 of the DFG Manual,including Cieanup Procedure Xll-6 (gel-chromatographic cleanup), has been used successfully in many laboratories because of its broad applicability for the gas-chromatographic determination of pesticide residues in foodstuffs. DFG method S19 is also included in the respective European Standards. Subsequently, a modification of the extraction and partition step has been implemented. The modified method requires less experimental effort and eliminates the use of dichloromethane, which is an undesirable solvent for toxicological and ecological reasons. As the results from validation studies demonstrate,... 

Traditional electrophoresis and capillary electrophoresis are competitive techniques as both can be used for the analysis of similar types of samples. On the other hand, whereas HPLC and GC are complementary techniques since they are generally applicable to different sample types, HPLC and CE are more competitive with each other since they are applicable to many of the same types of samples. Yet, they exhibit different selec-tivities and thus are very suitable for cross-validation studies. CE is well suited for analysis of both polar and nonpolar compounds, i.e. water-soluble and water-insoluble compounds. CE may separate compounds that have been traditionally difficult to handle by HPLC (e.g. polar substances, large molecules, limited size samples). 

Sandra and David [37] have reported on validation studies for SHS-HSGC, Py-HSGC, on-line LVI-GC, on-line SPME-GC, on- and off-line SPE-GC, on-line and off-line derivatisation-GC, SBSE-TD-GC, and PTV-LC(SEC)-GC. 

A theoretical approach has also been taken to predicting the droplet formation from a pMDI, and this was followed by experimental validation studies [31,32],... 

Details of the specific types of apparatus need not normally be given except for nonstandard processes. A flow chart of the manufacturing operation and the in-process controls (and acceptance limits) is required. Proposals for alternative processes will need to be supported by appropriate data to show that the finished products resulting from these are consistent with the finished product specification. Certain manufacturing operations such as mixing may require additional information on quality parameters monitored during production and prior to batch release. Appropriate quality parameters should be included in the finished product specification regardless of the outcome of validation studies (e.g., content uniformity for solid and semi-solid products). 

The sterilization processes described in the Ph Eur are preferred, especially terminal sterilization in the final container alternative processes have to be justified. All sterilization processes will need to be described and appropriate in-process controls and limits included. Where Ph Eur prescriptions are followed, there should be a statement to this effect in the application. Most of this information should be discussed in the development pharmaceutics section. Reference is made to the specific guidelines on ethylene oxide sterilization and irradiation sterilization, which are discussed further below. The possibility of parametric release for terminal processes such as saturated steam and irradiation is mentioned (see below). For all sterile products there should be a sterility requirement included in the finished product specification regardless of the outcome of validation studies. 

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