Correlation between acidic ninhydrin and HPLC methods to evaluate fraudulent addition of whey in milk

Abstract

-The acidic ninhydrin spectrophotometric method (ANSM) for quantitative determination of free and bound sialic acid of milk glycoprotein has been proved to be fast and efficient for routine detection of fraudulent addition of rennet whey to fluid milk. In this research the ANSM was compared with the high performance liquid chromatography (HPLC) method, internationally recommended for caseinomacropeptide (CMP) determination, which besides its high accuracy is more sophisticated and requires trained personnel. For several sample conditions (raw milk and milk with variable added amounts of rennet cheese whey), the methods showed an excellent linear correlation, with r = 0.981 when milk was deproteinized with a 120 g·L -1 final concentration of trichloroacetic acid (TCA) concentration. The best correlations could be seen with final concentrations of 100 g·L -1 and 80 g·L -1 TCA; respectively, r = 0.992 and 0.993. Milk / adulteration / whey / addition / acidic ninhydrin / HPLCRésumé -Détection de l'addition frauduleuse de lactosérum au lait de consommation : comparaison de la méthode à la ninhydrine acide (détermination de l'acide sialique) avec la méthode HPLC de détermination du CMP. La méthode spectrophotométrique à la ninhydrine acide (ANSM : acidic ninhydrin spectrophotometric method) pour la détermination des formes libres et liées de l'acide sialique s'est avérée être rapide et performante pour la détection en routine de l'addition frauduleuse de lactosérum doux dans le lait de consommation. Dans l'étude réalisée, l'ANSM a été comparée avec la méthode recommandée au niveau international, à savoir la méthode de détermination du caséinomacropeptide (CMP) par HPLC qui est certes très précise mais requiert un équipement sophistiqué et du personnel hautement qualifié. Une excellente corrélation linéaire a été trouvée entre les deux méthodes lors de l'analyse de filtrats TCA (acide trichloroacétique) issus d'échantillons de lait cru additionnés de quantités variables de lactosérum doux soumis préalable-ment à un traitement thermique. Selon la concentration finale en TCA : 120 g·L -1 , 100 g·L -1 , 80 g·L -1 , les valeurs de corrélation linéaire atteignaient respectivement 0,981, 0,992 et 0,993.Lait / fraude / lactosérum / addition / ninhydrine acide / HPLC * Corresponding author: salvador@fea.unicamp.br 502 S.P. Fukuda et al.

Comparação entre três métodos analíticos para determinação de soro em leite cru refrigerado

“…Fukuda et al (2004) sugerem que a CLAE, metodologia sofisticada de alta sensibilidade, precisão e acurácia, seja utilizada em caso dúbio e em contraprova, por ser um processo oneroso, demorado, trabalhoso e que requer mão de obra qualificada. Somam-se a isso o custo da manutenção e o tempo de vida útil das colunas cromatográficas.…”

“…Essa técnica é internacionalmente aceita para identificar e quantificar tanto o CMP (Fukuda et al, 2004) quanto o método qualitativo, utilizando-se o reativo de Ehrlich para a determinação qualitativa do ácido siálico (Brasil, 2006). Entretanto, essas técnicas são caras, difíceis de serem executadas e requerem mão de obra especializada.…”

“…Tal metodologia, de acordo com Fukuda et al (1996;2004), pode ser utilizada como substituta das demais na detecção da adição de soro em leite. Esses autores ressaltam que o teor de CMP ou de ácido siálico em leite e seus derivados tem atuado como um importante marcador das ações proteolíticas.…”

Caseinomacropeptide index in UHT whole milk stored under different conditions of temperature and time

“…As an alternative to the routine for detecting fraudulent addition of whey to milk, the spectrophotometric ninhydrin acid may be used due to its excellent correlation to the official method (r = 0.981) (Fukuda et al, 2004), as well as by ELISA (Dupont et al, 2007).…”

Free and total GMP (glycomacropeptide) contents of milk during bovine lactation

“…Recently, FUKUDA, RÖIG & PRATA [8] (unpublished data) showed an excellent correlation between the quantitative sialic acid method (acid ninhydrin) and HPLC for GMP (r = 0.996), permitting the widespread use of the spectrophotometric method, a simple, inexpensive and rapid method.…”