ࡱ> q`ZbbjbjqPqP ::&I , , , @ !!!8"\x"d@ Fh"^F#(n#n#n# ' ' 'EEEEEEE$HhyJLF], *&@&@**F n#n#2cF{/{/{/* 8n# 8n#E{/*E{/{/A  Bn#" Yr!/-ADyF0FAJ.J(B"J, B, ''|{/J(d( ' ' 'FF /p ' ' 'F****@ @ @ !@ @ @ !@ @ @   PROTOCOL OF QUALITY ASSURANCE AND ORGANIZATION OF PROFICIENCY TESTING IN DAIRY LABORATORy Neven Antunac1, Natasa Mikulec1, imun Zamberlin1, Iva Horvat1 and Jasmina Havranek11 University of Zagreb, Faculty of Agriculture, Dairy Science Department, Svetosimunska 25, 10000 Zagreb, Croatia  HYPERLINK "mailto:antunac@agr.hr" antunac@agr.hr,  HYPERLINK "mailto:nmikulec@agr.hr" nmikulec@agr.hr,  HYPERLINK "mailto:szamberlin@agr,hr" szamberlin@agr.hr,  HYPERLINK "mailto:ihorvat@agr.hr" ihorvat@agr.hr,  HYPERLINK "mailto:jhavranek@agr.hr" jhavranek@agr.hr Abstract:Quality assurance of the analytical results is an obligation of all analytical laboratories which tend to be credible. In order to obtain accuracy and precision of results each laboratory implements suitable programmes according to EN ISO/IEC 17025:2006. Since 2005, Reference Laboratory of the Dairy Science Department is accredited according to the above mentioned norm. Validation of analytical methods and verification of validation parameters are part of good laboratory practice. Quality of analytical results is assured by use of reference methods and certified calibration standards and by participation in international proficiency trials which are organized by dairy laboratories from Germany, Italy, France, Switzerland and Slovenia. The Reference Laboratory of Dairy Science Department has its own protocol of quality control which is based on analytical procedures and methods which are traceable, accurate, reliable and applicable for certain purpose. At the same time, our laboratory organizes proficiency testing for other dairy laboratories. Results of analyses are statistically tested and on the basis of z-score we estimate the successfulness of measurements. The aim of this paper is to demonstrate the organisation of proficiency testing (Ring control) of analytical results for milk fat, protein, lactose and somatic cells count in milk for other dairy laboratories in Croatia, Bosnia and Herzegovina and Macedonia. Key words:Quality assurance, proficiency testing, milk  INTRODUCTION Milk quality depends on the number of different parameters which must be determined. For each parameter there are often several different methods. Minor errors in laboratory routines or errors of interpretations are usually found in results when different methods are used for the same parameter. For that reason unification of techniques and application of standard methods for determination of milk components must be accepted by local dairy laboratories and the National Central Laboratory for Milk. Those standard methods for determining microbiological and chemical components are crucial for estimating the price of milk. Therefore, quality assurance becomes necessary to ensure that the analytical results of milk components are accurate and very similar from one laboratory to another which is accomplished by the calibration of instruments with traceable calibration standards and participation in proficiency tests [1]. The aim of this paper is to demonstrate the organisation of proficiency testing (Ring control) of results for milk fat, protein, lactose and somatic cells count in milk for other dairy laboratories in Croatia, Bosnia and Herzegovina and Macedonia. experimental Individual cows milk samples for the Ring test must be fresh. Their chemical components content must comply with an ordinary range, from minimum to maximum values, depending on lactation stage. The number of somatic cell count and bacteria count in milk samples should comply with the National Milk Quality Regulation [2]. Materials The Reference Laboratory of the Dairy Science Department prepares secondary reference material for the Ring control once a month and 20 to 25 high quality cows milk samples are needed for preparing Ring samples. The quantity of samples depends on the number of registered participants in Ring control. An average number of participants is 20, and therefore about 3 litres per high quality sample is needed. Procedures Samples of milk are analyzed by the infrared spectrometry method (using the instrument Milkoscan FT 120, according to ISO 9622:1999) [3] to determine the content of milk fat, protein and lactose, the fluoro-opto-electronic method is used to determine somatic cell count (using the instrument Fossomatic Minor, according to ISO 13366-2:2006) [4], whereas the flow citometry method is used to determine an overall bacteria count (using the instrument Bactoscan FC, according to ISO 21187:2004) [9]. Based upon obtained results, 7 samples are chosen for Ring control for milk fat, protein and lactose using IR instruments as well as 7 samples for somatic cell count instruments using the fluoro-opto-electronic method. Samples must be of high hygienic quality [2] and must comprise a specific content range. After that chosen samples are preserved by bronopol, and as soon as the preservative is dissolved, samples must be homogenized in the homogenisator. Then samples are portioned in bottles of approximately 100 millilitres. Each participant of Ring control is given one set which consists of 7 samples whereas the Reference Laboratory retains at least 3 to 5 same sets. These sets are used to determine reference values for the content of milk fat, protein and lactose (for Ring test, IR instruments) and somatic cell count (for Ring test, flouro-opto-electronic instruments). Reference values are determined on the basis of repeated sample measurements by the instrumental-routine methods (Milkoscan FT 120 and Fossomatic Minor) and the result accuracy is checked by the reference methods [5, 6, 7, 8, 10]. According to obtained reference values, when Ring control participants results are received, statistical analysis is made to compare those results with reference values obtained in the Reference Laboratory. Prepared milk samples are kept in the refrigerator (5 3C) until they are sent over to Ring control participants. In transportation of samples, a portable refrigerator (5 3C) is used and participants must keep the samples under the same conditions until further analyses. The validity of milk samples is 5 days from sampling. The analysis procedure is as following: first of all samples must be left until they gradually reach the room temperature, then they must be heated in a water bath until they reach 41-43C. After that the sample must be gently shaken and then analysed. RESULTS AND DISCUSSIONS For each laboratory involved in the inter laboratory trial, an annual report is made. It refers to how successful all participants have been in analyzing milk fat, protein and lactose (Figure 1) and somatic cell count (Figure 2). In the analytical report, z-score is presented for each parameter. The z-score should not exceed 2.  Figure 1 - Annual report of laboratory analyses of milk fat, protein and lactose content  Figure 2 - Annual report of laboratory analyses of somatic cell Statistical analysis The statistical analysis is made according to AOAC Protocol [11]. The first stage in producing a score from a result x is obtaining an estimate of the bias which is defined as: bias = x X where X is true value or in practice the best estimate of X and x is measured value. Next step is comparing the bias estimate with a target value for standard deviation that forms the criterion of performance. A z-score is formed from: z = (x-X) /  where  is the target value for standard deviation. If X and  are good estimates of the population mean and standard deviation, then z will be approximately normally distributed with a mean of zero and unit standard deviation. As z is standardized, it is comparable for all analytes and methods. Values of z are combined to give a composite score for a laboratory in one round of proficiency test (Figure 1, Figure 2). The z scores can therefore be interpreted as follows: [z] d" 2 result is satisfactory 2 <[z] < 3 result is questionable [z] e" 3 result is unsatisfactory CONCLUSIONS Involving analytical laboratories in Ring control, we can control their accuracy of measured results on the instruments which are used for determining the content of milk fat, protein, lactose and somatic cell count in milk. REFERENCES [1] ISO-IEC 17025: General requirements for the competence of testing and calibration laboratories, (2005). [2] Croatian National Milk Quality Regulation, Croatian Official Bulletin, No. 102, (2000). [3] ISO 9622: Whole milk determination of milk fat, protein and lactose content Guidance on the operation of mid-infrared instruments, (1999). [4] ISO 13366 2: Milk Enumeration of somatic cells Part 2 Guidance on the operation of fluoro opto electronic counters, (2006). [5] ISO 13366 - 1: Milk Enumeration of somatic cells Part 1 Microscopic method, (1997). [6] ISO 1211: Milk Determination of fat content Gravimetric method (Reference method), (1999). [7] ISO 8968 2: Milk Determination of nitrogen content Part 2: Block digestion method, (2001). 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