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DNA-based verification that gelatin-containing foods and cosmetics do not contain pork products has always been a challenge due to DNA damage and destruction during gelatin production. 

In this study (open access) the authors report that – by careful optimisation of conditions – they could successfully apply a “traditional” PCR test to the problem.

They describe DNA extraction, post-isolation DNA analysis, annealing temperature and primer concentration optimization, specificity assay, amplification efficiency trial, sensitivity test, repeatability examination, and marketed sample analysis.

They report that the developed method demonstrated good specificity under optimized conditions. It achieved a good amplification efficiency of 101.2% with an R² of 0.994. The real-time PCR technique had a limit of detection of 1,316 pg in the sensitivity examination and a coefficient of variation of 0.81% in the repeatability testing.

They tested 10 retail samples (five facial mask cosmetics, food additive gelatin powder, two marshmallow products, and two gummy candy products), reporting that all of the samples displayed no amplification and were thus considered not to contain porcine DNA, consistent with the manufacturers’ labels.

The authors conclude that their real-time PCR method meets the validation criteria for qualitative analysis, including specificity, amplification efficiency, sensitivity, and repeatability.

Undeclared lard in confectionary products is a significant concern for consumers in many parts of the world who avoid pork on religious grounds.

This study (GBP30 download fee) used gas chromatography with flame ionization detection (GC–FID) to measure fatty acids, then principal component analysis (PCA) to detect porcine fatty acid biomarkers in imported chocolates and biscuits.

The authors report that total fat content ranged from 11.5 to 32.5%, with palm kernel-based chocolates enriched in lauric (42–52%) and myristic acids (18–20%), while other chocolates were dominated by palmitic, stearic, and oleic acids. Biscuits contained high proportions of palmitic and oleic acids (> 75%).

PCA of the complete fatty acid dataset separated lard-adulterated samples.. Targeted PCA using porcine biomarkers palmitic-to-oleic acid ratio and eicosadienoic acid confirmed this clustering.

Calibration using simulated lard–palm oil mixtures (0–15% w/w; five replicates per level) enabled quantitative estimation of lard .

Photo by Pawel Czerwinski on Unsplash

This paper (purchase required) describes the development and validation of a highly sensitive lateral flow immunoassay (LFIA) test kit for detecting trace pork in meat products. A rapid bead-based sample extraction was developed (1–5 min) for the  biomarker (porcine IgG).  The authors report that total test time to result was 20 minutes. The reported detection limit was 0.001 % (w/w), which is 5–500 × more sensitive compared to current commercial lateral flow kits. The LFIA was validated with a range of meat and processed food products, confirming its high specificity to pork without cross-reactivity to other animal species or non-meat ingredients. Moreover, a long-term stability study confirmed that the LFIA maintained its performance at room temperature storage for 2 years.

Differentiating gelatin species is an analytical challenge because of a lack of intact DNA.  Most speciation methods therefore target the profile of proteins.  Proteins are difficult to analyse - they are too large to measure directly by techniques such as LC-MS, without  prior breaking down, and their folded structure is also an important diagnostic parameter.  This structure is disrupted by many of the sampling and extraction procedures used in analytical method. Analysis of mixed gelatins is particularly difficult.

This method (open access) used a new approach based on the interaction of ethanol with amino acids inside a protein. Ethanol can denature globular proteins by disrupting intraprotein hydrogen bonds due to hydrophobic interactions. However, when added to solutions having proteins with considerable number of α-helices, ethanol can stabilize the protein structure and prevent aggregation. The specific effects of ethanol on protein structure and function can vary depending on the protein's composition and environment.

Attenuated Total Reflectance-Fourier Transform Infrared (ATR-FTIR) spectroscopy was used to leverage ethanol's differential effects on gelatin's amide bands for quantifying pork gelatin contamination in bovine gelatin.

The authors report that the method showed a strong linear correlation between contamination levels and amide band transmission, with detection and quantification limits of 0.85 and 2.85 mg/100 mg (pork in bovine), respectively. It effectively identified pork gelatin in halal candy, with recovery rates from 50.05 % to 103.69 %.

A new meat species authentication method designed for Halal food verification has been developed by scientists at the University of Münster, using a  SCIEX QTRAP^® 6500 LC-MS/MS system. BusinessWire reports that the method that can detect multiple species simultaneously.

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