The BioLuminizerTMhas been optimized for detection of bioluminescence patterns on HPTLC plates. The most important features are:
High quantum efficiency allowing short exposure times.
16 bit dynamic range to detect small changes in intensity.
Resolution optimized to resolve all details on the HPTLC plate following treatment withVibrio fischeri.
No dry-out, the bacteria are kept moist and luminescent for hours.
Stable environment allowing long term and differential measurements.
Predictable bioluminescence activity for reproducible results.
The CAMAG BioLuminizerTMstands out from similar system with its very smart and compact setup enabling precise and convenient positioning of the HPTLC plate, easy cleaning and compatibility with use under field conditions.
Figure 103. Comparison of different substance concentrations with two reference tracks shown on the right side. Dark spots represent reduced or stopped bioluminescence and can thus indicate the presence of toxic substances
Figure 104. Bioluminescence response for different toxicity levels
201 8.8 Bioluminescence Measurements
9 GMP/GLP-Conforming Operations in TLC
The abbreviations “GMP” and “GLP” have already been used in various parts of this book. Both of these concepts originated in the United States. The fundamental rules for the preparation of medicaments and the guarantee of their quality lie behind the lettersGMP(Good Manufacturing Practice). The fundamental principles are summa- rized and explained in “EU Guidelines on Good Manufacturing Practice for Medica- ments” [130].
The ground rules ofGLP(Good Laboratory Practice) were laid down for the or- derly planning, performance and monitoring of laboratory tests, especially those aimed at detecting harmful effects of medicaments, chemicals and plant protection agents, and sometimes also those of cosmetics and food additives.
Where is work done in accordance with GMP and GLP?
The GMP rules and GLP rules are equivalent, but simply apply to different areas. It can generally be said that in pharmaceutical factories and their control laboratories the work is done in accordance with GMP principles, the so-called GQLP (Good Quality Laboratory Practice). This also applies to pharmacists who supply medica- ments that they prepare themselves. All laboratories which carry out toxicological in- vestigations of the effects of medicaments on animals, for which an application for per- mission from the German Institute for Medicaments and Medical Products (BfArM) must be made, require an official certificate stating that they are working in accor- dance with GLP. Failing this, their test results are not recognized [131]. Also for this reason, all TLC users who have to perform their work in accordance with these ground rules are recommended to read the “GLP Handbook for Laboratory Technicians”
[132].
The difference between routine laboratory work as performed today and that of 30 years ago lies not only in improved analytical methods and the benefits of electronic data processing. The main difference is the enormous paper mountain that represents the comprehensive documentation demanded by GMP/GLP and the frustration that sometimes goes with it.
The parameters that have to be documented according to the GMP/GLP rules can be expressed in the following short formula:
The burden of complying with these rules is sometimes relieved by witty colleagues who invent or pick up alternative meanings for the abbreviations, e.g. “GMP = gives more power, gives more paper, general minimization of productivity” etc. [133, 134].
However, humor can sometimes actually give a better understanding of these ideas.
For example, Wilhelm Busch has produced cartoons which illustrate the important ele- ments of GLP in picture form [131].
The history of GLP – whom have we to thank for all this?
The above question is often posed by young people starting their first job. It is there- fore appropriate at this point to quote an apt extract from an article on Good Labora- tory Practice by M. Schmidt [135]:
“In the early 1970s, the American Food and Drug Administration (FDA) discov- ered that, in animal experiments in the field of hazard identification (e.g. testing for acute and chronic toxicity, mutagenicity, teratogenicity, carcinogenicity), there was evi- dence of deceit and slovenly work when certification and registration were applied for.
Some testing laboratories had submitted data from a two-year study within a few weeks, and some had completed 2000 studies in one year. Also, experimental animals had become muddled up, although without any intention to mislead, and other mis- haps of a similar nature had also occurred, all of which placed the value of the results submitted in question.
The FDA reacted quickly by publishing the first draft of a GLP regulation in 1976.
The final version came into force in mid-1979. Since then, in laboratories in the United States, the data for the assessment of possible hazards to humans and the environment, when submitted for certification, permission, registration, or reporting for information purposes, must undergo regular inspections. If irregularities come to light, severe fines or even imprisonment could be imposed.
In 1981, the OECD (Organization for Economic Cooperation and Development) adopted the American GLP rules in their concept for the mutual acceptance of test data for the evaluation of chemicals.
In 1989, the FDA/OECD-GLP rules were incorporated intoEU law,thereby oblig- ing the member states to include these regulations in their national law. Since 1 August 1990, this EU requirement has been fulfilled by the Federal Republic of Germany, and the fundamental principles of GLP were incorporated into the chemical law (ChemG).”
From the above discussion, the following conclusion can be drawn:
Standard Operating Procedures (SOPs). In routine laboratory work, with its GMP/GLP rules, one is governed by a large number, often hundreds, of “SOPs”.
In these, clear and comprehensible descriptions ofall current routine processes are required (see, for example, the SOP “Registration, Printing and Archiving of Video Shots” in Section 8.4). M. Schmidt gives the following commentary on this [131]:
204 9 GMP/GLP-Conforming Operations in TLC
“The American origins of GLP can be clearly seen in the misconception that poor training of laboratory personnel can be compensated for by a flood of written instructions”. W. Günther comments as follows: “GLP gives the laboratory legal but not technical security” [136]. However, SOPs are a valuable basis for personal training! They are especially useful for revision if “he who always sees to it” is on holiday.
Because the terms operating procedure, standard operating procedure, testing pro- cedure, working procedure etc. are often used under various circumstances, the defini- tion of Standard Operating Procedures (SOPs) given by the Pharmaceutical-Technical Committee of the German Association of Pharmaceuticals Manufacturers (BAH) [137] is quoted here: “Standard Operating Procedures are documents that give infor- mation and instructions for the performance of operations that do not necessarily refer to one particular product or one particular task, e.g. the cleaning of equipment and plant, clothing to be worn in the production area, or climate control. They are essen- tiallygeneralinstructions of an organizational, administrative or technical nature and not specific instructions referring to specific products, manufacturing methods, clean- ing instructions etc.
Theproduct-specificrequirements and instructions are rather to be found in docu- ments such as manufacturing methods for the particular products, operating instruc- tions for particular machines and plant, cleaning and disinfection instructions for par- ticular equipment and plant etc.”
Having followed these necessary expositions on the theme of “GMP” and “GLP”, the question will occur to the user: “How can all these requirements be translated into practice in the area of TLC?”
Practical Tipsfor GMP/GLP-conforming operations in TLC:
Whenever possible, use coded plates.Laser-coded TLC and HPTLC plates preco- ated with silica gel 60 F254considerably simplify quality assurance in the laboratory.
Documentation and archiving of test results in accordance with GMP/GLP require- ments is then no longer dependent on the troublesome task of writing on the plates.
The name of the manufacturer, the item number, the charge number and the serial number of the plate are clearly visible and cannot be changed, being etched into the top edge of the plate (or on rotation through 90° on the left or right hand edge).
These markings cannot be tampered with and are automatically included in the photographic or video documentation of the chromatographic result [138].
205 9 GMP/GLP-Conforming Operations in TLC
Only use validated TLC methods in routine work.Validation in TLC is described in Section 9.1.
Use qualified/calibrated TLC equipment. The qualification/calibration of the equipment used is described in Section 9.2.
All the necessary parameters should be documented on raw data sheets.Suitable forms for GMP/GLP are given in Section 9.3.
Testing procedures, enabling work to be performed in a truly reproducible fashion, should be written as a result of method developments.This subject is discussed in Section 9.4.
What will the future bring?
The rules of GMP and GLP are naturally not fixed for all time. They undergo a contin- ual process of renewal and optimization aimed at saving time and resources and avoid- ing pointless investigations – especially those involving animals. The second Meeting of Experts to reexamine the principles of GLP in the OECD took place in June 1996 in Paris (80 experts from 26 nations representing industry and also official bodies re- sponsible for supervision and assessment). The resulting recommended changes could be put into effect in late 1997 at the earliest [139]. In the second German GMP Confer- ence in November 1996 in Frankfurt/Main an intensive dialog with representatives of the American Health Authority (the FDA) took place for the first time, and this is to be continued [140]. An important topic for the future will be the increased use of computer-supported systems. The recommendations of the project group “GLP and Electronic Data Processing” of the GLP study group of the German Chemical Indus- try Association have been published [141, 142]. The aim of the proposed method of working is to minimize the cost of validation of computerized systems as far as possible while at the same time fulfilling the requirements of GLP. In the field of pharmaceuti- cal manufacturing processes, a paper on the validation of computerized systems has also been published, which points out ways of fulfilling GMP requirements while at the same time using information obtained for an effective revalidation and potential improvement of the process [143].
206 9 GMP/GLP-Conforming Operations in TLC