Variation of levels of aflatoxin M1 in raw milk from different localities in the central areas of Punjab, Pakistan

doi:10.1016/j.foodcont.2007.12.002

Food Control

Volume 19, Issue 12, December 2008, Pages 1126-1129

https://doi.org/10.1016/j.foodcont.2007.12.002 Get rights and content

Abstract

The occurrence of aflatoxin M₁ (AFM₁) in samples of raw milk of buffaloes and cows from different localities in the central areas of Punjab, the province of Pakistan, was determined by using high-performance liquid chromatography (HPLC) with prior clean-up step applying immunoaffinity columns. The present study has been designed to find out the variation of levels of aflatoxin M₁ in raw milk of different localities. Feed has major role for prevalence of aflatoxin M₁ in milk and different feed regimen is being used in different areas. Total 480 milk samples were analyzed, among these 360 were of buffalo milk and 120 were of cow milk. The percentage of AFM₁ contamination in buffalo milk and cow milk was 42.5% and 52.5%, respectively. The mean value of AFM₁ was 0.027 μg L⁻¹ in buffaloes’ milk and was 0.044 μg L⁻¹ in cows’ milk. In both types of milk, level of AFM₁ concentration was higher in milk samples obtained from urban and semi-urban areas and it was minimal in milk from rural areas.

Introduction

Aflatoxins are a group of closely related hepatocarcenogenic bisdihydrofurano metabolites produced by certain species of Aspergillus especially by certain strains of Aspergillus flavus and Aspergillus parasiticus. The common aflatoxins are B₁, B₂, G₁, G₂ and M₁. They are named after their respective innate fluorescence properties. Aflatoxin B₁ (AFB₁) is the most commonly occurring and the strongest carcinogenic amongst the aflatoxins. After the AFB₁ has been ingested, it is bio-activated by cytochrome P₋450 to a genotoxic epoxide, a DNA reactive metabolite that forms N⁷ – guanine adducts and it is also able to bind with proteins (Cupid et al., 2004). Aflatoxin M₁, which is metabolite of AFB₁, is excreted in the milk of lactating animals and has toxic properties similar to AFB₁ (Steyn, 1995).

The aflatoxins were declared as human carcinogens in 1987 by the International Agency for Research on Cancer (IARC) and the classification was confirmed by re-evaluation in 1992. Later, on the demonstrated toxic and carcinogenic effects of AFM₁, the toxin initially classified by IARC as a Group 2B human carcinogen (IARC, 1993), has now moved to Group 1 (IARC, 2002).

High-performance liquid chromatography (HPLC) is commonly used technique in latest aflatoxin determination (Elgerbi et al., 2004, Nachtmann et al., 2007, van Egmond and Dragacci, 2001). Methodology for determination of aflatoxin M₁ in milk has been greatly improved in recent years with the application of immunoaffinity columns (IAC) which provide a combination of extraction and clean-up stages. With the advent of immunoaffinity columns, AOAC Official Method 2000.08 has come into force. This method is meant for aflatoxin M₁ in milk using IAC by liquid chromatography with Final Action 2004. The previous AOAC liquid chromatographic method for aflatoxin M₁ and M₂ in fluid milk was AOAC Official Method 986.16 with Final Action 1990 (Official Methods of Analysis of AOAC International, 2000, chap. 49).

Feed of lactating animals has substantial role in the occurrence of AFM₁ in milk. Depending on the local conditions and traditions, different feed regimen is used in different areas like urban and rural. The present study has been designed to observe the levels of aflatoxin M₁ in raw milk of buffaloes and cows from different localities of urban, semi-urban and rural in the central areas of the Punjab province of Pakistan. While determining AFM₁ in buffalo milk samples, another dimension that is the variation in AFM₁ concentration in milk with respect to herd-size was also taken into consideration.

Section snippets

Milk samples

Samples of raw milk of buffaloes (360) and cows (120) were procured by directly approaching the milking sites during January 2007 to March 2007. Area for milk collection was divided into three categories namely, urban, semi-urban and rural. The milk samples were stored in freezer compartment inside a refrigerator until these were analyzed for AFM₁. The milk samples were placed in a cooler with icepacks during transportation.

Chemicals and standards

Acetonitrile (HPLC grade) of Sigma–Aldrich (Steinheim, Germany) was

Results and discussions

In this study IACs have been applied along with HPLC. Immunoaffinity columns have been successfully used in the analysis of aflatoxins in food and feed during the last few years (Scott & Trucksess, 1997). Many researchers have used IAC in combination with HPLC (Gurbay et al., 2006, Tuinstra et al., 1993) for the analysis of aflatoxins. The IAC in combination with Fluorometer was applied by Chiavaro, Cacchioli, Berni, and Spotti (2005) for determination of aflatoxin B₁ and aflatoxin M₁ in pig

Acknowledgements

Authors are highly grateful to Higher Education Commission, Islamabad for funding this project. The cooperation for providing facility of HPLC analysis by the Directorate WTO-Quality Control Laboratory, University of Veterinary and Animal Sciences, Lahore is highly appreciated and laudable.

References (18)

B.C. Cupid et al.
The formation of AFB₁-macromolecular adducts in rat and human and dietary levels of exposure
Food Chemical Toxicology
(2004)
A. Gurbay et al.
Assessment of aflatoxin M₁ levels in milk in Ankara. Turkey
Food Control
(2006)
I. Hussain et al.
A study on contamination of aflatoxin M₁ in raw milk in the Punjab province of Pakistan
Food Control
(2008)
C. Nachtmann et al.
Regional monitoring plan regarding the presence of aflatoxin M₁ in pasteurized and UHT milk in Italy
Food Control
(2007)
M. Sassahara et al.
Aflatoxin occurrence in foodstuff supplied to dairy cattle and aflatoxin M₁ in raw milk in the North of Prana state
Food and Chemical Toxicology
(2005)
P.S. Steyn
Mycotoxins, general view, chemistry and structure
Toxicology letters
(1995)
AOAC Official Methods of Analysis
Aflatoxin M1. Natural toxins
E. Chiavaro et al.
Immunoaffinity clean-up and direct fluorescence measurement of aflatoxin B₁ and M₁ in pig liver: comparison with high-performance liquid chromatography determination
Food Additives and Contaminants
(2005)
S. Dragacci et al.
Immunoaffinity column cleanup with liquid chromatography for determination of aflatoxin M₁ in liquid milk: collaborative study
Journal of AOAC International
(2001)

There are more references available in the full text version of this article.

Cited by (55)

Aflatoxins posing threat to food safety and security in Pakistan: Call for a one health approach
2023, Food and Chemical Toxicology
Aflatoxins are among the most important mycotoxins due to their widespread occurrence and adverse impacts on humans and animals. These toxins and/or their metabolites cannot be destroyed with cooking or boiling methods. Therefore, consumption of aflatoxin-contaminated food may lead to impaired growth, compromised immunity, stomach and liver cancer, and acute toxicity. These adverse effects along with food wastage might have detrimental consequences on a country's economy. Several studies from Pakistan reported a high prevalence of aflatoxins in food and feed commodities (Range; milk = 0.6–99.4%, cereals, and grains = 0.38–41%, animal feed = 31–100%). Notably, Pakistan reported very high figures of impaired child growth—stunted 40.2%, wasted 17.7% and underweight 28.9%—that could be associated with the higher aflatoxin prevalence in food items. Importantly, high aflatoxins prevalence, i.e. 100%, 69% and 60.5%, in children has been reported in Pakistan. Food and feed are more prone to aflatoxin contamination due to Pakistan's hot and humid climate; however, limited awareness, inadequate policy framework, and weak implementation mechanisms are the major obstacles to effective control. This review will discuss aflatoxins prevalence, associated risk factors, adverse health effects, required regulatory regime, and effective control strategies adopting the One Health approach to ensure food safety and security.
Health risk assessment of aflatoxin M1 in infant formula milk in IR Iran
2020, Food and Chemical Toxicology
In this study, two accurate, precise, selective and sensitive methods were developed for determining aflatoxin M1 (AFM1) in infant formula milk using immunoaffinity column clean-up followed by high performance liquid chromatography (HPLC) with fluorescence detection. The validated methods were used for determination of AFM1 in 29 samples of 6 different infant formula milk brands and the risk of AFM1 in infants aged zero to 6 months old was assessed using cancer risk, Margin of Exposure (MOE) and Hazard Index (HI). Only one sample (3.4%) was contaminated with AFM1. Although the results showed that MOE values for the mean and median exposure to AFM1 was <10,000 in infants, the additional cancer risk due to mean and median exposure to AFM1 in infant <6 months were 0.00010 and 0.00012 additional cases per year per 10⁵ individuals, respectively, which indicates no health concern. In addition, HI values for the mean and median exposure to AFM1 for infants were quite below one which indicates no health concern. To the best of our knowledge, this is the first report on risk assessment of AFM1 in infant formula milk consumed by Iranian infants <6 months old, presenting a low risk for the evaluated groups.
Quality control of beverages for health safety: Starting from laboratory to the point-of-care detection techniques
2019, Quality Control in the Beverage Industry: Volume 17: The Science of Beverages
Beverages are a class of liquid food item intended for human consumption to satisfy thirst but play important roles in human culture. Therefore, beverage contamination or fraud is also very popular and always occur with frightening predictability. According to the literature, beverages adulteration includes addition of toxic substances, like melamine, formaldehyde, urea, pesticides, metal ions, and various microorganisms exceeding a threshold value. Besides this, use of nanoparticles in consumer products has also increased in recent era as contaminants. Novel analytical methods have been proposed for the determination of these contaminants to assure the food safety and avoid health risks to consumers. Herein, in this chapter, we have summarized all the progresses made in the quality control in the beverages field by dividing the major contaminants into four categories: organic, inorganic, nanoparticles, and microorganisms for an easy understanding. A critical discussion has also been given on the progress, failure, and future of these sensing/separation methods for real-time applications.
Risk assessment and exposure to levels of naturally occurring aflatoxins in some packaged cereals and cereal based foods consumed in Accra, Ghana
2019, Toxicology Reports
Citation Excerpt :
With regards to aflatoxins, published findings of Inan et al. [7] suggested that there are a little over 20 recognized aflatoxins. Nonetheless, the four major ones critically considered are B1 (AFB1), aflatoxin B2 (AFB2), aflatoxin G1 (AFG1), and aflatoxin G2 (AFG2) while aflatoxin M1 (AFM1) and M2 (AFM2) are regarded as minor and have been reported as being the hydroxylated metabolites of AFB1 and AFB2 [8,9]. Among the five (5) notable mycotoxins in food, aflatoxins (B1, B2, G1 and G2) are the most toxic.
Aflatoxins are toxic secondary metabolites of fungal origin that contaminate agricultural commodities before, during and after harvest periods. A total of fifty-three (53) different foods (27 rice brands, 20 cereal based food brands and 6 pasta brands) were randomly obtained from the market and assessed for their different aflatoxin constitution (AFB1, AFB2, AFG1 and AFG2) as well as the total levels of the aflatoxins using High Performance Liquid Chromatography (HPLC) method. For the rice grain category, RS4 recorded the highest aflatoxin quantities of 65.77, 19.27, 1.02 μg/kg for AFB1, AFB2, AFG1 respectively and a total of 86.06 μg/kg which significantly differed (p < 0.05) from the other brands of foods. For the cereal based food category, CBS11 recorded the greatest quantities of 35.46, 4.92, 3.39 and 0.32 μg/kg for AFB1, AFB2, AFG1 and AFG2 respectively and a total of 45.1 μg/kg. For the pasta category, PS1 recorded the greatest quantities of 0.94 and 0.85 μg/kg for AFB1 and AFB2 respectively. Total aflatoxin quantities detected in some foods were above the acceptable limits set by the European Union which makes them unsafe and dangerous for human consumption. Recorded Estimated Daily Intakes (EDI) and Hazard Indices (HI) values were in the range of 3.9 × 10^-3 – 0.899 and 3.9 × 10^-4 - 0.0899 respectively. The risk was low since HI values obtained were less than 1.
Aflatoxin M<inf>1</inf> Contamination in Milk and Dairy Products
2017, Nutrients in Dairy and their Implications for Health and Disease
Milk and dairy products contain numerous nutritional compounds that have been associated with human health. However, they may also be a source of hazardous chemical or biological contaminants that can be harmful for humans. Aflatoxin M₁ (AFM₁), i.e., a secondary metabolite of fungi, has been considered as one of the most important toxic compounds in milk and dairy products because of its high stability during heat processing such as pasteurization and ultrahigh-temperature (UHT) treatment. On the other hand, the serious human health hazards of AFM₁ such as hepatotoxic, immunosuppressive, mutagenic, and carcinogenic effects are well established. In the current review, toxicity, occurrence of AFM₁ in milk and different types of dairy products, international regulations, and the latest efficient methods for determination of the toxin are described, which may help in establishing and carrying out strict monitoring strategies.
Milk Adulteration
2017, Nutrients in Dairy and their Implications for Health and Disease
The nutrients of milk are required for the proper functioning, growth, and development of the human body. The extensive consumption of milk and dairy products in Pakistan makes these foodstuffs targets for potential adulteration with financial gains for unscrupulous producers. Unfortunately, due to the unorganized and nonregulated marketing system in Pakistan, the quality of milk is hardly maintained at consumer level. Review of the literature has revealed that milk available to local populations is heavily adulterated with a variety of chemicals including heavy metals and aflatoxins. Microbial populations in raw milk are usually also beyond acceptable levels. The current milk safety situation in Pakistan is a pressing challenge and will continue to have serious implications for the nation’s economic infrastructure if corrective measures are not taken in a timely manner.

View all citing articles on Scopus

View full text

Variation of levels of aflatoxin M1 in raw milk from different localities in the central areas of Punjab, Pakistan

Abstract

Introduction

Section snippets

Milk samples

Chemicals and standards

Results and discussions

Acknowledgements

Food Chemical Toxicology

Food Control

Food Control

Food Control

Food and Chemical Toxicology

Toxicology letters

Aflatoxin M1. Natural toxins

Immunoaffinity clean-up and direct fluorescence measurement of aflatoxin B1 and M1 in pig liver: comparison with high-performance liquid chromatography determination

Food Additives and Contaminants

Immunoaffinity column cleanup with liquid chromatography for determination of aflatoxin M1 in liquid milk: collaborative study

Journal of AOAC International

Variation of levels of aflatoxin M₁ in raw milk from different localities in the central areas of Punjab, Pakistan

Immunoaffinity clean-up and direct fluorescence measurement of aflatoxin B₁ and M₁ in pig liver: comparison with high-performance liquid chromatography determination

Immunoaffinity column cleanup with liquid chromatography for determination of aflatoxin M₁ in liquid milk: collaborative study