Contamination of some foodstuffs marketed in Italy by fatty acid esters of monochloropropanediols and glycidol

ABSTRACT

Fatty acid esters of 3-monochloropropanediol (3-MCPD), 2-MCPD and glycidol (Gly) are food-processing contaminants that cause concerns about possible adverse health effects. The present study evaluates the contamination levels of the three ester classes in 130 samples of foodstuffs marketed in Italy covering 10 food categories, namely margarines, oils, roasted coffee, breakfast cereals, salted crackers, cookies, infant biscuits, rusks, breads and potato crisps. The analytical method employed is a so-called indirect method that entails MCPD/glycidol cleavage from their esterified forms, cleanup, derivatisation and GC-MS analysis. The MCPDs and glycidol concentrations (from esters) were found to be equal or a little higher than the levels reported in previous studies conducted in other European countries and described in the literature. 3-MCPD was the predominant compound in all foodstuffs analysed with the exception for rusks where Gly levels were slightly higher. Considering the sum of MCPD and Gly esters, the most contaminated foodstuffs were seed oils, followed by margarines and cookies, whereas roasted coffee, bread, rusks, cornflakes and infant biscuits were less contaminated with MCPDs and Gly concentrations often below LOQ or LOD values. Refined olive oil, potato chips and salted crackers showed contamination levels intermediate between the two above groups. The results of this study also confirm that the use of palm oil as an ingredient or frying medium is an important cause of increase of the levels of MCPD and Gly esters, especially in salted crackers, rusks and potato crisps. Finally, the Italian intake of 3-MCPD due to the various foods analysed has been calculated and related to TDI. The MoE for Gly was also estimated.     

Updated occurrence of 3-monochloropropane-1,2-diol esters (3-MCPD) and glycidyl esters in infant formulas purchased in the United States between 2017 and 2019

ABSTRACT

Fatty acid esters of 3-monochloropropane-1,2-diol (3-MCPD) and glycidol are potentially carcinogenic and/or genotoxic processing contaminants that are formed during the process of edible oil refining. Because of their toxicological properties, the presence of these compounds in refined oils and foods containing these oils, particularly infant formula, poses a potential food safety concern. For this reason, recent research efforts have focussed on the development of methods for the analysis of MCPD and glycidyl esters in infant formula in order to estimate levels of exposure. This work presents occurrence data for 3-MCPD and glycidyl esters in 222 infant formulas purchased in the United States between December 2017 and January 2019. The results of this study show a wide range of contaminant concentrations across four different manufacturers, with average bound 3-MCPD concentrations ranging from 0.035 µg g^?1 to 0.63 µg g^?1 and average bound glycidol concentrations ranging from 0.019 µg g^?1 to 0.22 µg g^?1. The data suggest that manufacturers B and C source palm oil produced with mitigation measures, leading to reduced amounts of 3-MCPD and glycidyl esters in their infant formulas. Additionally, comparison with a previously published study in our laboratory of the occurrence of 3-MCPD and glycidyl esters in infant formula purchased in the U.S. between 2013 and 2016 revealed that, since 2016, contaminant concentrations have decreased in products produced by manufacturers A, B, and C, while contaminant amounts in formulas from manufacturer D have slightly increased.     

Toxicological assessment of 3-chloropropane-1,2-diol and glycidol fatty acid esters in food

Fatty acid esters of 3‐chloropropane‐1,2‐diol (3‐MCPD) and glycidol are a newly identified class of food process contaminants. They are widespread in refined vegetable oils and fats and have been detected in vegetable fat‐containing products, including infant formulas. There are no toxicological data available yet on the 3‐MCPD and glycidol esters, and the primary toxicological concern is based on the potential release of 3‐MCPD or glycidol from the parent esters by lipase‐catalyzed hydrolysis in the gastrointestinal tract. Although 3‐MCPD is assessed as a nongenotoxic carcinogen with a tolerable daily intake (TDI) of 2 μg/kg body weight (bw), glycidol is a known genotoxic carcinogen, which induces tumors in numerous organs of rodents. The initial exposure estimates, conducted by Federal Institute for Risk Assessment (BfR) under the assumption that 100% of the 3‐MPCD and glycidol are released from their esters, revealed especially that infants being fed commercial infant formula could ingest harmful amounts of 3‐MCPD and glycidol. However, the real oral bioavailability may be lower. As this gives rise for toxicological concern, the currently available toxicological data of 3‐MCPD and glycidol and their esters are summarized in this review and discussed with regard to data gaps and further research needs.     

Occurrence of 3-MCPD and glycidyl esters in edible oils in the United States

Fatty acid esters of 3-monochloropropanediol (3-MCPD) and glycidol are processing contaminants found in a wide range of edible oils. While both 3 MCPD and glycidol have toxicological properties that at present has concerns for food safety, the published occurrence data are limited. Occurrence information is presented for the concentrations of 3-MCPD and glycidyl esters in 116 retail and/or industrial edible oils and fats using LC-MS/MS analysis of intact esters. The concentrations for bound 3-MCPD ranged from below the limit of quantitation (<LOQ) to 0.09 mg kg^?1 (ppm) in 22 unrefined oils and from 0.005 to 7.2 mg kg^?1 (ppm) in 94 refined oils. The concentrations for bound glycidol ranged from <LOQ to 0.03 mg kg^?1 (ppm) in unrefined oil samples and from <LOQ to 10.5 mg kg^?1 (ppm) in processed oil samples. The highest concentrations for both 3-MCPD and glycidol were seen in refined palm oil and palm olein samples. Palm olein samples also contained a higher percentage of 3-MCPD in mono-ester form than any other type of oil.     

微生物油脂中氯丙醇酯的形成及应对措施综述

氯丙醇及其酯类是国际社会广泛关注的食品污染物之一,近年来发现氯丙醇酯广泛存在于各种食用油脂及含油脂食品中,而引起油脂行业的广泛关注。本文较为详细的总结了目前国内外对油脂中氯丙醇酯的毒性、检测方法、形成机制及控制方式等的研究现状,并对微生物油脂中可能的产生机制和应对措施进行了概述,以指导微生物油脂及食品的生产和消费,推动我国微生物油脂食品的健康发展。     

Development of an analytical method for the simultaneous analysis of MCPD esters and glycidyl esters in oil-based foodstuffs

Substantial progress has been recently made in the development and optimisation of analytical methods for the quantification of 2-MCPD, 3-MCPD and glycidyl esters in oils and fats, and there are a few methods currently available that allow a reliable quantification of these contaminants in bulk oils and fats. On the other hand, no standard method for the analysis of foodstuffs has yet been established. The aim of this study was the development and validation of a new method for the simultaneous quantification of 2-MCPD, 3-MCPD and glycidyl esters in oil-based food products. The developed protocol includes a first step of liquid–liquid extraction and purification of the lipophilic substances of the sample, followed by the application of a previously developed procedure based on acid transesterification, for the indirect quantification of these contaminants in oils and fats. The method validation was carried out on food products (fat-based spreads, creams, margarine, mayonnaise) manufactured in-house, in order to control the manufacturing process and account for any food matrix–analyte interactions (the sample spiking was carried out on the single components used for the formulations rather than the final products). The method showed good accuracy (the recoveries ranged from 97% to 106% for bound 3-MCPD and 2-MCPD and from 88% to 115% for bound glycidol) and sensitivity (the LOD was 0.04 and 0.05 mg kg^?1 for bound MCPD and glycidol, respectively). Repeatability and reproducibility were satisfactory (RSD below 2% and 5%, respectively) for all analytes. The levels of salts and surface-active compounds in the formulation were found to have no impact on the accuracy and the other parameters of the method.     

Estimated US infant exposures to 3-MCPD esters and glycidyl esters from consumption of infant formula

A dietary exposure assessment was conducted for 3-monochloropropane-1,2-diol (3-MCPD) esters (3-MCPDE) and glycidyl esters (GE) in infant formulas available for consumption in the United States. 3-MCPDE and GE are food contaminants generated during the deodorisation of refined edible oils, which are used in infant formulas and other foods. 3-MCPDE and GE are of potential toxicological concern because these compounds are metabolised to free 3-MCPD and free glycidol in rodents and may have the same metabolic fate in humans. Free 3-MCPD and free glycidol have been found to cause adverse effects in rodents. Dietary exposures to 3-MCPDE and GE from consumption of infant formulas are of particular interest because formulas are the sole or primary food source for some infants. In this analysis, US Food and Drug Administration data on 3-MCPDE and GE concentrations (as 3-MCPD and glycidol equivalents, respectively) in a small convenience sample of infant formulas were used to estimate exposures from consumption of formula by infants 0–6 months of age. 3-MCPDE and GE exposures based on mean concentrations in all formulas were estimated at 7–10 and 2 µg/kg bw/day, respectively. Estimated mean exposures from consumption of formulas produced by individual manufacturers ranged from 1 to 14 µg/kg bw/day for 3-MCPDE and from 1 to 3 µg/kg for GE.     

油脂中3-氯丙二醇酯形成的化学反应机制

3-氯丙二醇酯是油脂及油脂食品在热加工过程中生成的副产物,目前被认为是潜在的食品安全危害因子之一.本文介绍了油脂中3-氯丙二醇酯形成的3种化学反应机制,普遍认为是氯离子的亲核进攻导致了3-氯丙二醇酯的形成.其中,第1种机制认为氯离子亲核反应的中间体环酰氧鎓离子是3-氯丙二醇酯的前体;第2种机制认为是亲核的氯离子直接攻击甘油骨架上的酯基或质子化羟基;第3种机制认为缩水甘油酯可能作为氯离子亲核反应的中间体而成为其前体.     

Change of fatty acid esters of MCPD and glycidol during restaurant deep frying of fish nuggets and their correlations with total polar compounds

This study examined the fatty acid esters of 2-monochloropropane-1,2-diol (2-MCPD), 3-monochloropropane-1,2-diol (3-MCPD) and glycidyl esters (GEs) in frying oils during fish nuggets deep frying. The 3-MCPD esters significantly increased in the first 12 h and then decreased, whereas 2-MCPD esters increased to a maximum at 24 h (0.69–0.81 mg kg⁻¹) and then decreased. The GEs decreased with frying time and degraded to approximately 0.05 mg kg⁻¹ after 36 h in all frying oils. The correlation results showed that the 3-MCPD esters had a positive correlation with 2-MCPD esters (R = 0.910), so 3-MCPD esters could be an indicator of the presence of MCPD esters in frying oils. Strong correlations were found between MCPD esters and the peroxide values and p-anisidine values, indicating that changes in MCPD esters may be related to oil oxidation. These results may improve our understanding of MCPD esters and GEs in frying oils and find ways to control them during frying.     

Glycidyl Fatty Acid Esters in Refined Edible Oils: A Review on Formation,Occurrence, Analysis,and Elimination Methods

Glycidyl fatty acid esters (GEs), one of the main contaminants in processed oils, are mainly formed during the deodorization step in the refining process of edible oils and therefore occur in almost all refined edible oils. GEs are potential carcinogens, due to the fact that they readily hydrolyze into the free form glycidol in the gastrointestinal tract, which has been found to induce tumors in various rat tissues. Furthermore, glycidol has already been identified as a “possible human carcinogen’’ (group 2A) by the Intl. Agency for Research on Cancer (IARC). Therefore, significant effort has been devoted to inhibit and eliminate the formation of GEs. The aim of this review is to provide a comprehensive summary on the following topics: (i) GE occurrence data for different edible oils and oil‐based food products, (ii) precursors of GEs, (iii) factors influencing the formation of GEs, (iv) potential reaction mechanisms involving the leaving group and reaction intermediates, and (v) analytical methods, including the indirect and direct methods. More importantly, the various elimination methods for GEs in refined edible oils are being reviewed with focus on 3 aspects: (i) inhibition and removal of reactants, (ii) modification of reactive conditions, and (iii) elimination of GE products.     

食用油中3-氯丙醇酯的研究进展

氯丙醇酯污染是近年来国际上出现的食品安全热点问题之一,尤其是3-氯丙醇酯污染更为突出,各种食用油或油脂食品中都含有一定量的3-氯丙醇酯。总结了国内外对3-氯丙醇酯的研究现状,介绍了3-氯丙醇酯来源、毒理和安全性、可能的形成机制、检测方法、污染水平、控制措施和精炼过程中形成的影响因素等方面的内容,并提出了目前急需解决的问题。旨在推动我国油脂食品行业的健康发展。     

Application of gastrointestinal modelling to the study of the digestion and transformation of dietary glycidyl esters

Glycidyl esters (GEs) are known to be formed during vegetable oil processing. Because of their structure, it has been hypothesised that GEs, like fatty acid esters of chloropropanols (MCPD esters), may be accepted as substrates by gut lipases to release the epoxide glycidol. If confirmed such a hypothesis would be important for risk assessment since glycidol is considered as a genotoxic carcinogen. In the present study, biotransformation was investigated using static and dynamic gastrointestinal models. During the experiments, aliquots were analysed for non-digested GEs using liquid chromatography-time-of-flight-mass spectrometry (LC-ToF-MS). In the static model, a fast hydrolysis of GEs was observed as a result of lipase action. Lipase was very efficient at pH 4.8, and totally inhibited at very low pH (1.7). In the absence of lipase, GEs were found to be relatively stable. The potential impact of food matrix was studied using milk in a dynamic model simulating human physiological conditions. The fast, pH-dependent hydrolysis of GEs was further confirmed. The possible transformation of the digestion products was then investigated using gas chromatography coupled to mass spectrometry (GC-MS), mainly the epoxide ring-opening to glycerol followed by additional reactions. In any conditions applied, neither 2- nor 3-mono-chloropropanediol (2- nor 3-MCPD) were formed, indicating that a ring-opening of the epoxide group of GEs or glycidol followed by a reaction with chloride was unlikely. A small transformation of glycidol into glycerol was observed after longer incubation time correlated with a low pH. This suggested that ring-opening and reaction with water is possible in strongly acidic conditions. Overall, it is concluded that GEs are rapidly digested by gut lipases to form glycidol. Consequently, GEs should be considered as sources of glycidol exposure. In addition, risk assessment of GEs can likely rely on hazard identification and characterisation data specific for glycidol.     

Formation mechanisms of monochloropropanediol (MCPD) fatty acid diesters in refined palm (Elaeis guineensis) oil and related fractions

Monochloropropanediol (MCPD) fatty acid esters are process contaminants generated during the deodorisation of edible oils. In particular, MCPD diesters are found in higher abundance in refined palm oil than other edible oils. In the present study, a series of model reactions mimicking palm oil deodorisation has been conducted with pure acylglycerols in the presence or absence of either organic or inorganic chlorine-containing compounds. Results showed that the bulk of MCPD diesters are formed above 200°C through the reaction of organochlorines with triacylglycerols (TAG). Additional experiments confirmed that this reaction can be initiated during palm oil deodorisation by hydrogen chloride (HCl) gas evolved through the thermal degradation of organochlorines present in the oil. Therein, the majority of the ultimately produced MCPD diesters are the result of HCl reacting with TAG, via protonation, followed by the elimination of a fatty acid residue. Two possible MCPD diester formation mechanisms are highlighted, both of which involve acyloxonium ion reactive intermediates. Investigations with pure TAG regio-isomers showed that MCPD ester formation is regioselective and the sn-1(3) position of the glycerol backbone is favoured.     

Mitigation Strategies for the Reduction of 2‐ and 3‐MCPD Esters and Glycidyl Esters in the Vegetable Oil Processing Industry

The refining of vegetable oils leads to the formation of 2‐ and 3‐monochloropropane‐1,2‐diol esters (2‐ and 3‐MCPD‐E), and glycidyl esters (Gly‐E). A literature review was performed aiming to provide up‐to‐date knowledge on mitigation strategies during oil refining that can reduce the formation of these three processing contaminants. The review used the database Scopus and covered the period from 2009 to 2017. Most of the 18 papers dealt with palm oil and two papers with vegetable oil. Most studies focused on 3‐MCPD‐E, some on Gly‐E, and none on 2‐MCPD‐E. Water degumming was able to reduce the concentrations of 3‐MCPD‐E by 84% and Gly‐E by 26%. Neutralization of the oil reduced concentrations of 3‐MCPD‐E by 81% and Gly‐E by 84%. Bleaching with synthetic magnesium silicate reduced the 3‐MCPD‐E concentration by 67%. For the deodorization step, several mitigation strategies, such as double‐deodorization, the addition of various antioxidants, or a longer deodorization time, can reduce the formations of 3‐MCPD‐E by 82% and Gly‐E by 78%. Postrefining mitigation, including the use of absorbents, enzymes, or rebleaching of the oil, has also been reported to produce desirable contaminant reduction. Postrefining treatment with calcinated zeolite was able to reduce the 3‐MCPD‐E concentration by 19% and the Gly‐E concentration by 77%. Applying combined mitigation strategies to multiple steps of oil refining is likely crucial in order to adequately reduce levels of 3‐MCPD‐E and Gly‐E.     

婴配食品中氯丙醇酯的污染来源及控制措施

氯丙醇酯类污染物主要形成于植物油精炼过程的脱臭工序,在食品加工过程中其含量也可能会变化。不同种类的油脂中污染物含量水平不同,不同工艺生产的同种油脂中污染水平也有较大差异,通过改善生产工艺可在较大程度上减少该污染物的含量。精炼植物油是婴配食品中脂肪的重要来源,在迚行产品配方设计时,不仅应考虑使其脂肪及脂肪酸在含量与结构上与母乳匹配,还应考虑可能引入的氯丙醇酯类污染物含量水平,尽可能从源头上减少此类污染物的带入。本文通过介绍氯丙醇酯的形成机制、婴配食品中脂肪来源及生产工艺及氯丙醇酯类污染物的主要类型,以确定该污染物的来源,幵提出可采取的延缓措施,从而帮助生产企业从理论和实践水平上控制此类污染物的暴露水平。     

食用油中缩水甘油酯的检测方法研究进展

缩水甘油酯(GEs)痕量存在于精炼食用油中。食用油中缩水甘油酯检测方法主要包括间接检测法和直接检测法。间接检测法即将缩水甘油酯通过酯交换反应,解离出缩水甘油,再通过衍生化后进行GC-MS测定。直接检测法在不破坏缩水甘油酯结构的基础上,先经过或不经过样品净化步骤直接测定。对食用油中缩水甘油酯检测方法的样品处理过程、仪器条件、结果进行综述,并对检测方法的优缺点进行分析。     

Mass-defect filtering of isotope signatures to reveal the source of chlorinated palm oil contaminants

This paper reports new insights at the molecular level into the route of a worldwide problem of the food industry: the occurrence of monochloro-propanediol (MCPD) esters. The application of mass defect-driven workflows is described to generate a hypothesis on the identity and occurrence of those thermally labile, chlorinated contaminant precursors that may act as chlorine donors during the formation of MCPD esters. For the first time, holistic mass-defect filtering of isotope signatures is used to pinpoint completely unknown and unexpected chlorine-containing substances naturally present in various extracts of palm fruit and partially and fully refined oils. Supervised multivariate analysis showed the effective classification of samples from various stages of industrial processing, suggesting that these steps strongly impact a complex and dynamic pool of chlorinated substances. In-vitro experiments confirmed that several of these naturally occurring chlorinated plant constituents decompose upon heat treatment, thus potentially being a source of chlorine for further reactions with palm oil lipids in a subsequent chlorination cascade. It is hypothesised that during oil refining the organochlorines naturally present in palm fruits act as a ‘chlorine source’ for the generation MCPD diesters. This discovery implies that industrial efforts targeting the mitigation of chlorinated substances must intervene at the earliest possible production stage or preferably even prior to oil processing. Current performance and limitations of mass-defect filtering are discussed and future developments are outlined.     

Washing bleached palm oil to reduce monochloropropanediols and glycidyl esters

The formation of toxic compounds, potentially carcinogenic, during food processing has been considered an important food safety issue. Among them, particular attention has been given to 3-monochloropropane-1,2-diol esters (3-MCPDE), 2-monochloropropane-1,3-diol esters (2-MCPDE) and glycidyl esters (GE), which can be formed during vegetable oil refining, especially palm oil. These substances may pose a health risk to humans due to their toxicity and carcinogenicity. The aim of this study was to investigate the effect of washing bleached palm oil (BPO) with different solvents, and evaluate the reduction of 3-MCPDE, 2-MCPDE and GE as well as assess the quality parameters of the final product. For this purpose, we used two types of washing with different solvents. A single washing was carried out in one step and a double washing in two steps using a solvent gradient. Single washing had a limited reduction in the levels of 3-MCPDE and 2-MCPDE and resulted in an increased level of GE, whereas double washing slightly reduced 3-MCPDE and 2-MCPDE and resulted in a significant reduction of GE levels. The reduction achieved in this study was up to 17.1% for 3-MCPDE, 56.4% for 2-MCPDE and 76.9% for GE levels. The reduction of 3-MCPDE and 2-MCPDE might be due to the removal of part of the ethanol-soluble chlorinated precursors from the oil which suggests that highly lipophilic forms of these substances are present in BPO. The substantial reduction on GE levels might be associated with the removal of the precursors present in the oil such as diacylglycerols. Thus, the washing treatment could be used as a supplementary strategy to reduce processing contaminants from palm oil, especially GEs.     

Verification and evaluation of monochloropropanediol (MCPD) esters and glycidyl esters in palm oil products of different regions in Malaysia

ABSTRACT

This study was conducted to investigate on the effect of different sampling regions of palm-refined oils and fats on the 2- and 3-monochloropropanediol fatty acid esters (MCPDE) and glycidol fatty acid esters (GE) levels. The American Oil Chemists’ Society (AOCS) Official Method Cd 29a-13 on the determination of MCPDE and GE in edible oils and fats by acid transesterification was successfully verified and optimised, with slight modification using 7890A Agilent GC system equipped with 5975C quadrupole detector. The determined limits of detection (LOD) for MCPDE were 0.02 mg kg^-1 and 0.05 mg kg^-1 for GE. The method performance has showed good recovery between 80% and 120% for all pertinent compounds with seven replicates assayed in three separate days. Round robin test with two European laboratories, i.e. Eurofins and SGS, has shown compliance results with those of the present study. Among the sampling regions, only one refinery located in the central region of Malaysia showed a significant increment of the MCPDE and GE levels after refining process. The GE level averaging at 2.5 mg kg^-1 was slightly higher than that of 3-MCPDE averaging at 1.3 mg kg^-1. Both esters were preferentially partitioned into the liquid phase rather than the solid phase after fractionation. However, the overall results exhibited no direct correlation between the esters content and the different sampling locations of the palm oil products in Malaysia. Analysis of total chlorine content also displayed significant variations between sampling locations which clearly show its effect on the chlorine content in the CPO samples.     

Factors impacting the formation of Monochloropropanediol (MCPD) fatty acid diesters during palm (Elaeis guineensis) oil production

Recently, organic and inorganic chlorinated compounds were detected in crude and commercially refined palm oils. Further, the predominant formation mechanism of monochloropropanediol (MCPD) diesters at high temperatures (>170–180°C) was revealed. The present study involved the development and comparison of solutions to mitigate MCPD diester levels in oils from various stages of palm oil production. Partially refined palm oil samples and oil extracted from fresh palm fruits were submitted to bench-top deodorisation experiments. Application of glycerol and ethanol as refining aids during the deodorisation of refined-bleached palm oil proved to be moderately effective; about 25%–35% reduction of MCPD diester levels was achieved. Washing crude palm oil with ethanol–water (1:1) prior to deodorisation was also an effective strategy yielding an ～30% reduction of MCPD diester contents. Washing palm fruit pulp before oil extraction, however, was most impactful, resulting in a 95% reduction of MCPD diesters when compared to the deodorised control oil. This suggests that intervention upstream in the process chain is most efficient in reducing levels of these contaminants in refined oils. Following the study, a root-cause analysis was performed in order to map the parameters potentially responsible for the occurrence of MCPD diesters in refined palm oil and related fractions.