Critical Evaluation of Two Qualitative Analytical Approaches for Multiclass Determination of Veterinary Drugs in Bovine Muscle Using UHPLC-Q-Orbitrap: The Wind of Change in Brazilian Monitoring.

Food safety is recognized as a main requirement for consumers, food industries, and official laboratories. Here, we present the optimization and screening qualitative validation of two multianalyte methods in bovine muscle tissues by ultra-high-performance liquid chromatography coupled to high-resolution mass spectrometry with an Orbitrap-type analyzer, operated with a heated ionization source in positive and negative mode. This aims for not only the simultaneous detection of veterinary drugs regulated in Brazil but also the prospection of antimicrobials not yet monitored. Two different sample preparation procedures were applied: method A-generic solid-liquid extraction with 0.1% formic acid (v/v) in an aqueous solution of EDTA 0.1% (w/v)-acetonitrile-methanol (1:1:1, v/v/v), followed by an additional ultrasound-assisted extraction and method B-QuEChERS. In both procedures, selectivity showed satisfactory conformity. From a detection capability (CCß) equivalent to ½ the maximum residue limit, >34% of the analyte resulted in a false positive rate of <5%, preponderant by the QuEChERS method, which exhibited a higher yield of the sample. The results showed the potential application of both procedures in the routine analysis of foods by official laboratories, enabling the expansion of this methodological portfolio as well as its analytical scopes, thus optimizing the control of residues of veterinary drugs in the country.

Determining performance parameters in qualitative multivariate methods using probability of detection (POD) curves. Case study: Two common milk adulterants.

A strategy for determining performance parameters of two-class multivariate qualitative methods was proposed. As case study, multivariate classification methods based on mid-infrared (MIR) spectroscopy coupled with the soft independent modelling of class analogy (SIMCA) technique for detection of hydrogen peroxide and formaldehyde in milk were developed. From the outputs (positive/negative/inconclusive) of the samples, which were unadulterated and adulterated at target value, the main performance parameters were obtained. Sensitivity and specificity values for the unadulterated and adulterated classes were satisfactory. Inconclusive ratios 12% and 21%, respectively, for hydrogen peroxide and formaldehyde were obtained. To evaluate the performance parameters related to concentration, Probability of Detection (POD) curves were established, estimating the decision limit, the capacity of detection and the unreliability region. When inconclusive outputs were obtained, two additional concentration limits were defined: the decision limit with inconclusive outputs and the detection capability with inconclusive outputs. The POD curves showed that for concentrations below 3.7gL-1 of hydrogen peroxide and close to zero of formaldehyde, the chance of giving a positive output (adulterated sample) was lower than 5%. For concentrations at or above 11.3gL-1 of hydrogen peroxide and 10mgL-1 of formaldehyde, the probability of giving a negative output was also lower than 5%.

Detection of several common adulterants in raw milk by MID-infrared spectroscopy and one-class and multi-class multivariate strategies.

A sequential strategy was proposed to detect adulterants in milk using a mid-infrared spectroscopy and soft independent modelling of class analogy technique. Models were set with low target levels of adulterations including formaldehyde (0.074g.L-1), hydrogen peroxide (21.0g.L-1), bicarbonate (4.0g.L-1), carbonate (4.0g.L-1), chloride (5.0g.L-1), citrate (6.5g.L-1), hydroxide (4.0g.L-1), hypochlorite (0.2g.L-1), starch (5.0g.L-1), sucrose (5.4g.L-1) and water (150g.L-1). In the first step, a one-class model was developed with unadulterated samples, providing 93.1% sensitivity. Four poorly assigned adulterants were discarded for the following step (multi-class modelling). Then, in the second step, a multi-class model, which considered unadulterated and formaldehyde-, hydrogen peroxide-, citrate-, hydroxide- and starch-adulterated samples was implemented, providing 82% correct classifications, 17% inconclusive classifications and 1% misclassifications. The proposed strategy was considered efficient as a screening approach since it would reduce the number of samples subjected to confirmatory analysis, time, costs and errors.

An appropriate and systematized procedure for validating qualitative methods: its application in the detection of sulfonamide residues in raw milk.

The lack of well-established references for the validation of qualitative analyses and the increasing demand for reliable binary responses were the main motivating factors for this study. A detailed procedure for single-laboratory validation of qualitative methods is proposed. The experimental design and the tools for data analysis were based on the theoretical background, as well as the aspects of efficiency, convenience and simplicity. Four experimental steps were defined, as follows: (i) preliminary tests for the determination of the concentration range, (ii) a study of the rates, unreliability region, detection limit, and the accordance and concordance values, (iii) a study of the selectivity in the presence of known interferences, and (iv) a study of robustness. The applicability of the procedure was demonstrated by the validation of a qualitative commercial kit for detecting sulfonamide residues in raw milk using both the visual and instrumental reading techniques. Reliability rates of 100% were obtained for the blank samples. For the samples spiked with sulfamethazine at 10.8 and 108 µg L(-1) and with sulfadimethoxine or sulfathiazole at 10 and 100 µg L(-1), the reliability rates ranged from 93.3 to 100%. Selectivity was demonstrated using trimethoprim as a potential interferent. The method was considered robust for the factors of the temperature (54 and 58°C) and time (6 and 10 min) for incubating the test strips. The estimated detection limits and unreliability regions confirmed the suitability of the kit for this purpose, based on the legislated residue limits.

Tendências em validação de métodos de ensaios qualitativos / Trends in implementing the validation of qualitative methods of analysis

A crescente importância das análises qualitativas e a necessidade de confiabilidade dos resultados analíticos para subsidiar as tomadas de decisões nos diversos setores da cadeia produtiva de alimentos são inquestionáveis, acarretando impactos na saúde pública, na economia e nos direitos do consumidor. Contudo, a validação desses métodos tem sido um ponto crítico na implantação de sistemas de gestão da qualidade e nos processos de acreditação de laboratórios. Apesar da existência de protocolos bem estabelecidos para validação de métodos quantitativos, ainda existe uma lacuna no desenvolvimento de abordagens para a implementação da metrologia em análises qualitativas. Neste contexto, o presente trabalho apresenta uma discussão sobre o tema validação de métodos qualitativos, abrangendo definições, delineamento experimental e análise de dados para avaliação dos parâmetros de desempenho aplicáveis como taxas de falsos resultados, taxa de confiabilidade, taxa de seletividade, taxa de sensibilidade, limite de detecção, região de perda de confiabilidade, acordância, concordância e robustez.