The manifestation of aflatoxins in feed and food is a major issue in the world as its presence leads to some health problems. This study investigates the incidence of aflatoxin M₁ (AFM₁) contamination in raw milk samples which were collected from Punjab, Pakistan. The Cluster Random Sampling technique was used to collect 960 milk samples from five different regions, and samples were collected every month. The AFM₁ level in raw milk was analyzed by the ELISA technique. The findings demonstrate that 70% of samples exceeded the United States permissible maximum residue limits (MRL 0.50 µg/L), with an overall AFM₁ level that ranged from 0.3 to 1.0 µg/L. AFM₁ contamination varied with the season: The highest average contamination was detected in winter (0.875 µg/L), followed by autumn (0.751 µg/L), spring (0.654 µg/L), and summer (0.455 µg/L). The Eastern region exhibited the highest average AFM₁ contamination (0.705 µg/L). Milk samples from the Northern region were found to be widely contaminated, as 86.9% samples exceeded the US MRL, followed by the Eastern region, with 72.3% samples being contaminated with >0.5 µg/L AFM₁. The study indicated that the raw milk supply chain was heavily contaminated. Recommendations and remedial measures need to be developed by regulatory authorities to improve the raw milk quality.

(1) Background: Multiple attempts have been conducted to correlate milk keeping quality with chemical, physical or bacteriological parameters. These methods only measure the chemical changes in milk produced by bacteria. Headspace solid-phase micro-extraction (HS-SPME) is an economic and recent method used to measure both volatile compounds and microbial load in milk, also allowing to keep the quality of the milk product. (2) Methods: The present study was conducted to identify and measure the off-flavoring volatile compounds through gas chromatography coupled with flame ionization detector (GC-FID) and the microbial load of pasteurized fluid milk stored at different temperatures, as a possible indicator of its keeping quality. (3) Results: The highest results were obtained to acetone, followed by butanone, pentanal and ethanol. These mean values were significantly enhanced from the 0 to 19th day of storage, at 10 °C. At day 19th, the minimum score for aroma, flavor and overall acceptability were also recorded as 4.33 ± 0.17, 4.02 ± 0.06, 4.00 ± 0.04, respectively. Likewise, maximum values for standard plate count (Log₁₀ CFU 15.54 ± 0.40 mL⁻¹) and total psychotroph count (Log₁₀ CFU 11.67 ± 0.30mL⁻¹) were reported at 10 °C and 4 °C. (4) Conclusion: HS-SPME/GC-FID methodology revealed to be very sensitive and capable to be applied in volatile compounds quantification in pasteurized milk produced during the storage period at different temperatures.