Analytica Chimica Acta 406 (2000) 257–278 Validation of bias in multianalyte determination methods. Application to RP-HPLC derivatizing methodologies Àngel Mart´ ınez a,∗ , Jordi Riu a , Olga Busto b , Josep Guasch b , F. Xavier Rius a a Departament de Qu´ ımica Anal´ ıtica i Qu´ ımica Orgànica, Facultat de Qu´ ımica, Universitat Rovira i Virgili, Pl. Imperial Tarraco, 1, E-43005 Tarragona, Catalonia, Spain b Departament de Qu´ ımica Anal´ ıtica i Qu´ ımica Orgànica, Unitat d’Enologia del CeRTA, Universitat Rovira i Virgili, c/Ramón y Cajal, 70, E-43005 Tarragona, Catalonia, Spain Received 16 September 1998; received in revised form 9 September 1999; accepted 24 September 1999 Abstract This paper reports a new approach for validating bias in analytical methods that provide simultaneous results on multiple analytes. The validation process is based on a linear regression technique taking into account errors in both axes. The validation approach is used to individually compare two different chromatographic methods with a reference one. Each of the two methods to be tested is applied on a different set of data composed of two real data sets each. In addition, three different kinds of simulated data sets were used. All three methods are based on RP-high-performance liquid chromatography (HPLC) and are used to quantify eight biogenic amines in wine. The two methods to be tested use different derivatizing procedures: precolumn 6-aminoquinolyl-n-hydroxysuccinimidyl carbamate (AQC) and oncolumn o-phtalaldehyde (OPA), respectively. On the other hand, the reference method uses derivatization with OPA precolumn. Various analytes are determined in a set of samples using each of the methods to be tested and their results are regressed independently against the results of the reference method. Bias is detected in the methods to be tested by applying the joint confidence interval test to the slope and the intercept of the regression line which takes into account uncertainties in the two methods being compared. The conclusions about the trueness of the two methods being tested varied according to whether the joint confidence interval test was applied to data obtained from various biogenic amines considered simultaneously or individually. ©2000 Elsevier Science B.V. All rights reserved. Keywords: HPLC; Biogenic amines; Method validation; Linear regression; Joint confidence interval 1. Introduction Biogenic amines need to be determined in fer- mented beverages because they are potentially toxic when consumed in large amounts [1]. Many methods for quantifying the biogenic amine content in food ∗ Corresponding author. Tel.: +34-977558187; fax: +34-977559563. E-mail address: martinez@quimica.urv.es (A. Mart´ ınez). have been described (based on i.e. gas chromatog- raphy [2,3] and high-performance liquid chromato- graphic (HPLC) techniques [4–6]). However, proce- dures based on RP-HPLC have commonly been used as the amines can be injected automatically [7], even without previous treatment of the samples. Most of the RP-HPLC analytical methods used to determine biogenic amines are based on derivatization reactions which improve the selectivity and sensitivity of the different procedures. 0003-2670/00/$ – see front matter ©2000 Elsevier Science B.V. All rights reserved. PII:S0003-2670(99)00766-7