Detection of Pork in Beef Meatballs Using LC-HRMS Based Untargeted Metabolomics and Chemometrics for Halal Authentication

Abstract

Adulteration of high-quality meat products using lower-priced meats, such as pork, is a crucial issue that could harm consumers. The consumption of pork is strictly forbidden in certain religions, such as Islam and Judaism. Therefore, the objective of this research was to develop untargeted metabolomics using liquid chromatography-high resolution mass spectrometry (LC-HRMS) combined with chemometrics for analysis of pork in beef meatballs for halal authentication. We investigated the use of non-targeted LC-HRMS as a method to detect such food adulteration. As a proof of concept using six technical replicates of pooled samples from beef and pork meat, we could show that metabolomics using LC-HRMS could be used for high-throughput screening of metabolites in meatballs made from beef and pork. Chemometrics of principal component analysis (PCA) was successfully used to differentiate beef meatballs and pork meatball samples. Partial least square-discriminant analysis (PLS-DA) clearly discriminated between halal and non-halal beef meatball samples with 100% accuracy. Orthogonal projection to latent structures-discriminant analysis (OPLS-DA) perfectly discriminated and classified meatballs made from beef, pork, and a mixture of beef-pork with a good level of fitness (R2X = 0.88, R2Y = 0.71) and good predictivity (Q2 = 0.55). Partial least square (PLS) and orthogonal PLS (OPLS) were successfully applied to predict the concentration of pork present in beef meatballs with high accuracy (R2 = 0.99) and high precision. Thirty-five potential metabolite markers were identified through VIP (variable important for projections) analysis. Metabolites of 1-(1Z-hexadecenyl)-sn-glycero-3-phosphocholine, acetyl-l-carnitine, dl-carnitine, anserine, hypoxanthine, linoleic acid, and prolylleucine had important roles for predicting pork in beef meatballs through S-line plot analysis. It can be concluded that a combination of untargeted metabolomics using LC-HRMS and chemometrics is promising to be developed as a standard analytical method for halal authentication of highly processed meat products.

Phytochemical analysis, antioxidant, α-glucosidase inhibitory activity, and toxicity evaluation of Orthosiphon stamineus leaf extract

“…Samples were prepared following Windarsih et al 19 with slight modifications. Analysis was performed by adding LC–MS-grade methanol into O. stamineus leaf extracts (water and ethanol extract).…”

Analysis of Pork in Beef Sausages Using LC-Orbitrap HRMS Untargeted Metabolomics Combined with Chemometrics for Halal Authentication Study

“…It has also been utilized to investigate the changes of lipid metabolites in Hengshan goat sausages during preservative treatment [ 32 ]. In addition, LC–HRMS metabolomics and chemometrics have been used for detection of pork in raw beef meat [ 22 ] and tuna meat [ 33 ], and the presence of pork in beef meatballs has been successfully detected by LC–HRMS non-targeted metabolomics and chemometrics [ 34 ].…”

Identification of volatile compounds in several meat and bone broth using Solid Phase Micro Extraction-Gas Chromatography Mass Spectrometry (SPME-GCMS) for initial detection of Halal and Non-Halal Food

“…The increase of Muslim population give rise to the demand for halal foods. Based on the Islamic shariah, pork is a non-halal meat that is prohibited from being consumed by Muslim [1]. However, due to economic reasons the adulteration of meat and meat product with lowerpriced pork meat has become a global issue.…”

“…However, those techniques require rigorous sample preparation, costly reagents, and is time-consuming. Numerous instrumental methods have also been reported for meat products authentication based on the volatile compounds in pork [6][7][8], beef and pork [9][10][11][12][13], lamb [14][15][16][17][18][19], goat [20], and chicken [1,[21][22].…”

“…The result showed that NCB contained more complex volatiles and exhibited a richer aromatic profile compared with BB. NCB contained 2,4-decadienal, 2,4-decadienal, 2,4-nonadienal, 2-nonenal as the aromatic compound with the highest flavor dilution (FD) factor in both broths [1] [21]. Ten key favor substances, such as 1-phellandrene, linalool, myrcene, limonene, ocimene, anethole, terpineol, dipropyl disulfide and 2,4-decadienal, were investigated [22].…”