I.V. Soma Raju      

Cross contamination is the real concern of the cleaning  and validation-articles" class="alinks-link" title="pharmaceutical cleaning validation">cleaning validation, need to be addressed  for the  effectiveness of the procedure being used. The effectiveness of cleaning shall be proven using  appropriate sampling techniques, analytical methods and the evaluation of potential contamination .

Of course any of the procedure shall be validated  against to its intended application. Several questions need to be addressed before validating the cleaning process and also it is necessary to think that the objective of cleaning process shall be considered the whole activities of the application in terms of objective, activities and the conclusion. While making the conclusion it shall be reviewed for any inherence associated obviously . In the process of thinking about the possible inherences of analytical methods, had a look on how the same sample can allow the different assessments based on the  procedures  adopted which is the sprout for this article .

Before going to the actual chapter ,recap of the basic facts  of the concept. As we know there are two types of samplings are involved in the evaluation of cleaning process that have been found acceptable. The most desirable is the direct method of sampling the surface of the equipment(Swab) according to FDA’s clean pot concept. Another method is the use of rinse solutions.

Swab sampling is a crucial step in sample preparation. It is not possible to obtain as such sample like rinse samples, it involves some level of dilution to collect and prepare the swab sample to make convenient for the analysis. The actual swab sample is the contaminant on the  specified surface. But it is not possible to analyze the contaminant from the surface directly. It is to be extracted using the best possible and practical ways. Generally 10 fold dilution of swab sample is essential and is evident when we observe the procedure.

Procedure :

1. Pre-treat the swabs in the sample solvent, and squeeze the swabs
2. Swab the specified surface in an uni-direction firmly to cover the entire area.
3. put the swab in to a test tube and add10 ml of sample solvent (extractable solvent)  and sonicate to extract the residue by sonication. (Firms will choose different folds to dilute , generally 10 to 30 )
4. Filter the extract sample and analyze the sample using the respective analytical method.

All these days we are considering that the swab technique is invasive in terms of sampling but not other ways, but It has been observed from the general practices of evaluation of contamination from swab samples due to the LOQ of the analytical method used .

Let us discuss the most common practices in the industry and how they are effecting from the actual scenario in making wrong diversions due to the  perception of the analytical procedure with out consideration of method’s performance level and  the practices adopted.(Scope of the article)

Giving the platform to think over the consequences with a possible cases and  their  risk assessment to explain how the same sample will allow  the different assessments  and the conclusion for better possible ways based on the evaluation procedures adopted and analytical techniques involved.

                     

Practice- 1: Practice of  multiplying the test results with respective to the dilutions made to the swab sample while calculating the content of residue .

Procedure:

 Calculation:                                                                          Au

                              Content of residue in ppm           =   ---------------------- x C  x 10

                                                                                   As x recovery factor

Where Au and As are responses of sample and standard solution respectively and C is the concentration of standard solution in ppm and 10 is the dilution factor of swab sample.

                     Weight of standard (g) x 10ml  x 1000000

C (ppm) = -------------------------------------------------------------

                   100 (stock dilutionin ml)x 100ml (std dilution in ml)

Details of the swab sample and the methods are:

Method’s Limit of detection is 0.5 ppm

Method Range is 1.5 ppm (LOQ) to  30 ppm

Example : The result of different samples :  for sample 1 : not detected,

                                                               sample 2 : 0.6  ppm

                                                               for sample 3 : 18 ppm

Other parameters:  As appropriate based on the analytical technique.

Inference of  practice-1

Evaluation	Sample-1 (Result: Not detected)	Sample-2  (Result: 0.6ppm( below      LOQ of the method LOQ=1.5 ppm))	Sample-3 (Result: 1.8 ppm)
	Even though the swab result is not detected , it is possible the actual contamination can be 5PPm, because the level of contamination from the swab sample will be measured as 0.5 ppm (below LOD of the method) instead of 5ppm due to the inherence of the sample 10 folds dilution), obviously the result will be not detected .	Even though the swab result is 0.6ppm ,it is possible the actual contamination can be 6 ppm, because the level of contamination from the swab sample will be measured as 0.6 ppm ( below LOQ of the method) instead of 6 ppm due to the inherence of the sample (10 folds dilution)preparation, obviously the result will be error one from the actual  and is made the sample as below LOQ level while measuring .	Though the level of contamination from this swab sample will be measured as 1.8 ppm ( above the LOQ) due to the inherence of the sample (10 folds dilution) , The reproducibility of the results may accurate , because the practice of multiplying the results as per the sample dilution will not be effected when the measurements  (concentrations)are found with in the range of method .
Risk assessment	From the evaluation of Case-1 it is evident that the approach of multiplying the swab results according to the dilutions made for the swab samples is not recommended and is to be noticed , that  it is justifiable like the swab samples -3 only  (when the level of contamination is with in the range even after 10 folds dilution) and it could not be practical to decide the accuracy of the result based on the level of contaminants rather than their potential.
Recommen-    dation	Shall not be adopted

Practice -2 :  Practice of  injecting more quantity of sample than the standard to compensate the dilution with respective to the dilution of swab preparation .

Procedure:

Standard preparation  :   10 ppm solution of respective analyte (contaminant)

Collection of  swab solution : Swab the surface on a predetermined  sampling point using swab stick and immediately transfer the swab in to a test tube and  add 10 ml of solvent (using pipette) and sonicate for about 2 minutes

Instrument conditions :

Injection volume of standard: 5 µl

Injection volume of swab sample: 50 µl

Other parameters: As appropriate based on the analytical technique.

In case of TLC methods : by visual observation of intensities only.

Incase of HPLC,GC , TOC and any other:

Calculate the content of residue  in ppm taken by the formula

                                          Au

  Content (ppm) =   --------------------------- x C

                                As x recovery factor

Where Au and As are the responces of sample and standard solution respectively and C is the concentration of standard solution in ppm

    Weight of standard (g) x 10ml x 1000000

C (ppm) = -------------------------------------------------------------

                   100 (stock dilutionin ml)x 100ml (std dilution in ml)

Inference of practice -2

Evaluation	When we inject sample 50µl(10 times higher than the standard(if standard injection volume is 5µl )  the responses of both  will be similar, since the correction of the sample dilution was made inherently (physically) which will allow the exactness to the real scenario(actual results )  .
Risk assessment	It is not  possible with all type of techniques ( like UV, titremetry),  possible like GC,,HPLC,TLC where the injection volumes involved in the method. Though needs to evaluate the impact  on result considering the similarities between standard and sample peak shape(controlling peak symetry and correlation of intensities (in case of tlc)  etc .
Recommendation	Shall  be adopted  for the analytical techniques having provision of  injecting the samples in a quantitative manner assuring the validated methods.

Practice-3 :  Standard addition method influencing the requirement of method ‘s range & recovery .

Detailed below giving a typical example to interpret the case to pursue in a practical  approach of this kind of practice.

Procedure:

Standard preparation  :  Weigh accurately about 10 mg of standard substance in a 100 ml of volumetric flask.  Dissolve and dilute to volume with methanol (stock). Further dilute 10.0 ml of  this solution to 100 ml with  methanol .

Collection of  swab solution : Swab the surface on a predetermined  sampling point using swab stick and immediately transfer the swab in to a test tube and  add 9.5 ml of methanol (using pipette), sonicate for about 2 minutes .

Preparation of  swab sample solution: Pipette out 0.5 ml of standard stock solution  in a 5 ml volumetric flask, dissolve and dilute to volume with swab solution and sonicate for about 2 minutes.

Instrument conditions :

Injection volume of standard: 20 µl

Injection volume of swab sample: 20 µl

Other parameters: As appropriate based on the analytical technique.

Procedure:

Calculate the content of residue  in ppm taken by the formula

                           Au                    

 Content (ppm) =  ------------ x standard dilutions x sample dilution10 (swab sample /dilution factor)

              As x recovery factor

Where Au is corrected absorbance of sample and  As is the absorbance of standard solution (10ppm) respectively and C is the concentration of standard solution in ppm

Corrected response of sample solution (Au) =  response absorbance of swab

sample-  response  obtained from standard solution (AS)

recovery factor : Methods’s recovery obtained from   individual surface recovery studies of method validation ( example: When the swab method recovery is 88% the recovery factor is 0.88) 

Note : Should be considered the result (with out any further calculation) as below limit of detection when Au value has got either negative or zero.

 

                       Weight of standard (g) x 10ml x 1000000

C (ppm) = --------------------------------------------------------

                         100 (stock dilutionin ml)x 100ml (std dilution in ml)

Inference of practice -3

Evaluation	For example a surface contains 12 ppm of contaminant and got diluted to 10times due to the swab sampling procedure, then the level of contaminant in swab preparation will be 1.2 ppm which is below LOQ of the method ( objective of the article) and is appropriate after adding the specified concentration to measure the proper concentration(11.2ppm which is above LOQ) because of the spiking of 10 ppm standard solution  to the swab preparation, and finally subtracting the added amount from the swab preparation results will be obtain the actual result of swab . Hense this  methodology is giving the platform to bring the sample concentration is always with in the range of method at the time of its measurement .
Risk assessment	Using the standard addition method  will give a solution to over come the analytical issues of swab samples due to its dilution against to the methods LOQ concern and also need to assure the  method’s accuracy be fore adopting. It should be noticed that this methodology cannot be possible for TLC method .
Recommendation	Can be adopted in most of the cases and is one of the better option to over come the objective of the issue.

Summary:

 It is not unusual to see firms use extensive sampling and testing programs following the cleaning process without ever really evaluating the impact of evaluation on effectiveness of the procedure being used.

Cleaning validation is a scientific demonstration of the effectiveness of cleaning.  We are making our efforts to achieve the maximum possible effectiveness using the proper protocols ,sampling techniques, SOP’s, validated analytical methods ,but overlooking the inherent issue of the method’s performance level .The proper choice of method and methodology will help to prevent the obvious errors in conclusion of the effectiveness of cleaning validation. 

About Author:

Soma raju,D.G.M.

Quality control . Hetero Labs limited , Hyderabad, AP-India.

Soma raju has more than 12 years experience in validation and compliance in the indian pharmaceutical industry and achieved the credential ,continuing top-1 of all the countries(world wide) for the topic Pharmaceutical Manufacturing .US, offered by Brain bench US certification .

Mailing address: I.V.Soma raju , 12-228, Jaya Towers , F.No:102,  Adarsh nagar, Opp-IDPL colony, Hyderabad-37, AP.

Mail I.D : ivsraju@gmail.com , somaraju@heterodrugs.com Phone numbers : mobile 91-9396243315,  Res: 91-40-65917918