Current Sample Preparation Methods and Determination Techniques for the Determination of Phthalic Acid Ester Plasticizers in Edible Oils.

In the process of production, processing, transportation, and storage of edible oils, the oils inevitably come into contact with plastic products. As a result, plasticizers migrate into edible oils, are harmful to human health, and can exhibit reproductive toxicity. Therefore, the determination of plasticizers in edible oils is very important, and a series of sample preparation methods and determination techniques have been developed for the determination of plasticizers in edible oils. Phthalic acid ester (PAE) plasticizers are the most widely used among all plasticizers. This review aims to provide a comprehensive overview of the sample preparation methods and detection techniques reported for the determination of PAEs in edible oils since 2010, focusing on sample preparation methods of edible oils combined with various separation-based analytical techniques, such as gas chromatography (GC) and liquid chromatography (LC) with different detectors. Furthermore, the advantages, disadvantages, and limitations of these techniques as well as the prospective future developments are also discussed.

Isolation of Two Plasticizers, Bis(2-ethylhexyl) Terephthalate and Bis(2-ethylhexyl) Phthalate, from Capparis spinosa L. Leaves.

Many plants have been known to be contaminated and accumulate plasticizers from the environment, including water sources, soil, and atmosphere. Plasticizers are used to confer elasticity and flexibility to various fiber and plastic products. Consumption of plasticizers can lead to many adverse effects on human health, including reproductive and developmental toxicity, endocrine disruption, and cancer. Herein, we report for the first time that two plasticizers, bis(2-ethylhexyl) terephthalate (DEHT) and bis(2-ethylhexyl) phthalate (DEHP), have been isolated from the leaves of Capparis spinosa L. (the caper bush), a plant that is widely used in food seasonings and traditional medicine. 297 mg/kg of DEHT and 48 mg/kg of DEHP were isolated from dried and grounded C. spinosa L. leaves using column chromatography and semi-preparative high-performance liquid chromatography. Our study adds to the increase in the detection of plasticizers in our food and medicinal plants and to the alarming concern about their potential adverse effects on human health.

Occurrence and distribution of phthalate esters and microplastics in wastewater treatment plants in Taiwan and their toxicological risks.

Phthalate esters (PAEs) are fat soluble synthetic chemicals, usually regarded as plasticizers for being added in numerous plastic products. Thus, environmental and health hazards of PAEs are associated with increasing plastic pollution. In this study, PAEs from sludge samples collected from water, sewage, and industrial treatment plants (N = 17) were analyzed using Gas Chromatography/Mass Spectrometry. Microplastics (MPs) were also quantified and correlated with PAEs. Results showed the highest average PAE concentrations in sewage treatment plants. The greatest ΣPAEs concentration were found in sewage treatment plant (STP4) with 32,414 µg/kg dw, while the lowest found in water treatment plant (WTP3) with 2062 µg/kg dw. Among different PAEs, di-(2-ethyl hexyl) phthalate (DEHP) contributes the highest. Similarly, DEHP, di-n-octyl phthalate (DnOP) and diisononyl phthalate (DiNP) significantly correlated with the total PAEs indicating their large contribution to sludge contamination. The abundance of microplastics in sludge ranged between 1 and 7 MP/g, highest at ITP6, but not detected in some stations. While microplastics may potentially increase PAEs, there was no significant relationship between ΣPAEs and MP abundance. The estimated human daily intake of DEHP and di-n-butyl phthalate (DnBP) when contaminated sludge be used showed low toxicological risks to exposed adults. This research presents the sludge characteristics, PAEs, and microplastic concentrations in different wastewater treatment plants in Taiwan. PAE contamination was highly contributed by domestic and industrial wastes shown by their significant amounts in STP and ITP. Results further provide evidence for potential sludge recycling (WTP sludge) and application to soil.

Persistent organic pollutants (PCBs and PCDD/Fs), PAHs, and plasticizers in spices, herbs, and tea - A review of chromatographic methods from the last decade (2010-2020).

Edible and highly demanded plant-derived products such as herbs, spices, and tea may be subjected to exogenous contamination of well-known chemical hazards such as persistent organic pollutants (POPs), and emerging ones such as plasticizers, affecting negatively the safety of these food commodities. This fact has led to the increasing analysis of exogenous compounds including priority POPs such as polychlorinated dibenzo-p-dioxins (PCDDs) and polychlorinated dibenzofurans (PCDFs), and polychlorinated biphenyls (PCBs), as well as highly persistent polycyclic aromatic hydrocarbons (PAHs). Currently, plasticizer residues are also considered an emerging issue because of the extensive use in food packaging and potential migration into foodstuffs. In this review, the studies published from 2010 to 2020 were discussed, including the main extraction methods applied for these contaminants from herbs, spices, and tea, and it was revealed the trend toward the use of less solvent-consuming and time-effective methods. Chromatographic methods were also described, which were mainly combined with detection techniques such as classical or mass spectrometry (MS) detection. Finally, a comprehensive overview of the occurrence of these selected exogenous compounds was presented in the studied matrices, showing that their monitoring should be further investigated to ensure food safety of highly consumed condiments and tea.

Di(2-ethylhexyl) adipate plasticizer triggers hepatic, brain, and cardiac injury in rats: Mitigating effect of Peganum harmala oil.

Di(2-ethylhexyl) adipate (DEHA) is a widely used plasticizer and prevalent environmental contaminant. In this study, DEHA concentrations in the milk, cheese, and butter samples wrapped with food-grade commercial polyethylene films and stored at 4 °C for 30 days were detected using gas chromatographic analysis. Also, the effects of exposure to a high dose of DEHA for a long duration on the liver, brain, and heart of Wistar rats were assessed. Besides, the possible beneficial effect of Peganum harmala oil (PGO), in relieving DEHA induced adverse effects was explored. For this purpose, four groups (8 rats/group) were orally given physiological saline, PGO (320 mg/kg bwt), DEHA (2000 mg/kg bwt), or PGO + DEHA for 60 days. The results revealed that the DEHA concentrations in the tested dairy products were ordered as follows: (butter > cheese > milk). Notably, the detected levels in butter were higher than the specific migration limit in foods. DEHA induced a significant increase in the serum levels of glucose, alanine transaminase, aspartate transaminase, acetylcholine esterase, creatine kinase-myocardium bound, malondialdehyde, tumor necrosis factor-α, and interleukin-1ß. But, significant hypoproteinemia, hypoalbuminemia, hypoglobulinemia, and hypocholesterolemia were evident following DEHA exposure. A significant reduction in the serum level of superoxide dismutase, reduced glutathione, and brain-derived neurotrophic factor was recorded. Besides, a significant downregulation in hepatic CYP2E1, brain glial fibrillary acidic protein, and cardiac troponin I gene expression was noticed. Moreover, DEHA exposure induced a significant decrease in Bcl-2 immunolabeling, but Caspase-3 immunoexpression was increased. On the contrary, PGO significantly recused DEHA injurious impacts. Therefore, PGO could represent a promising agent for preventing DEHA-induced hepatotoxicity, neurotoxicity, and cardiotoxicity.

Cumulative risk assessment of phthalates in edible vegetable oil consumed by Chinese residents.

BACKGROUND: Phthalates have been widely used as plasticizers in various industries and are widely focused on in the international community as a result of their reproductive toxicity. Exposure of Chinese residents to phthalates via edible vegetable oil occurs often. In the present study, gas chromatography-mass spectrometry was used to detect the two main phthalates bis(2-ethylhexyl) phthalate (DEHP) and dibutyl phthalate (DBP) in four major edible vegetable oil sources: an edible oil blend, soybean oil, peanut oil and rapeseed oil (a total of 1016 samples), as collected throughout China. Furthermore, cumulative risk assessment was used to estimate the reproductive health risk to Chinese residents caused by the phthalates that come from edible vegetable oils. RESULTS: Both phthalates were detected in four major edible vegetable oil sources. The phthalate with the highest detection rate was DBP (13.48%), followed by DEHP (7.78%). The results of the cumulative risk assessment showed that the hazard indices of these two phthalates in edible vegetable oils were less than 1, except in soybean oil. Nevertheless, the two phthalates had the lowest detection rates in soybean oil, which were 1.94% (DEHP) and 5.16% (DBP). In China, contamination levels of phthalates in the soils where oil crops are cultivated have a great influence on the phthalate concentrations in edible vegetable oils. CONCLUSION: It is recommended that Chinese residents who are consuming soybean oil choose well-known brands and regularly change their brand of consumption. The phthalates in edible vegetable oils pose a relatively small reproductive health risk to Chinese residents. © 2019 Society of Chemical Industry.

Indoor Air Pollution in Cars: An Update on Novel Insights.

From a global viewpoint, a lot of time is spent within the indoor air compartment of vehicles. A German study on mobility has revealed that, on average, people spend 45 minutes per day inside vehicles. In recent years the number of cars has increased to around 43 million vehicles in private households. This means that more than one car can be used in every household. The ratio has been growing, especially in eastern Germany and rural areas. "Overall and especially outside the cities, the car remains by far number one mode of transport, especially in terms of mileage". Therefore, numerous international studies have addressed different aspects of indoor air hygiene, in the past years. In this paper, meaningful original studies on car indoor air pollution, related to VOCs, COx, PMs, microbials, BFRs, OPFRs, cigarettes, electronic smoking devices, high molecular weight plasticizer, and NOx are summarized in the form of a review. This present review aimed to summarize recently published studies in this important field of environmental medicine and points to the need for further studies with special recommendations for optimizing the interior air hygiene.

The use of of inulin, maltitol and lecithin as fat replacers and plasticizers in a model reduced-fat mozzarella cheese-like product.

BACKGROUND: Mono-, di- and oligosaccharides, polyhydric alcohols and lipids are three main types of plasticizers used to process food materials. In the present study, inulin, maltitol and lecithin were selected as representative oligosaccharide, polyhydric alcohol and lipid fat replacers, respectively. Their effects on the physicochemical properties of reduced-fat mozzarella cheese were evaluated. RESULTS: Lecithin reduced the hardness and increased the degree of free oil released. Inulin and lecithin decreased the hydrophobic interaction of reduced-fat cheese. Maltitol improved the elasticity of the reduced-fat cheese and increased the hydrophobic interaction within the casein matrix. Maltitol-added cheese had a lower glass transition temperature (Tg ) than the other cheeses. Maltitol significantly improved the stretchability of the reduced-fat cheese. CONCLUSION: The results obtained in the present study suggest that maltitol is an effective fat replacer in reduced-fat mozzarella cheese and might enhance the cheese's functional properties. The Tg of cheese was related to the water and fat content, fat replacer addition and cross-linking degree of casein. The relationship between Tg and the physicochemical properties of cheese will be studied in further research. © 2019 Society of Chemical Industry.

Dermal exposure to plasticizers in nail polishes: An alternative major exposure pathway of phosphorus-based compounds.

Phosphorus-based compounds are used as plasticizers in the manufacture of many products found in the indoor environment. Here we quantitatively investigated dermal exposure to phosphorus-based compounds contained in 45 nail polishes purchased in Japan. The alternative plasticizer triphenyl phosphate (TPhP) was detected in some samples of the nail polishes made in the USA (concentration, 1.1-1.8 wt%). The potential dermal exposure rates for TPhP, estimated using ConsExpo (version 5.0; Dutch National Institute for Public Health and the Environment), were in the range 200 (5%ile)-1700 (50%ile)-5000 (95%ile) ng kg-bw-1 day-1, which is more than 1400 times the reported values for exposure via dust ingestion and inhalation. Thus, dermal exposure via nail polish may be a major route of exposure to TPhP. The margin of exposure range for TPhP was 3.6 × 105-4.1 × 104-1.4 × 104. For comparison, the potential dermal exposure rate range for the conventional plasticizer dibutyl phthalate and the alternative plasticizer acetyl tributyl citrate was 360-3500-14,000 and 430-4100-17,000 ng kg-bw-1 day-1, respectively, and the margin of exposure range was 4.1 × 103-4.2 × 102-1.1 × 102 and 2.3 × 105-2.4 × 104-5.9 × 103, respectively.

Biodegradation patterns of the endocrine disrupting pollutant di(2-ethyl hexyl) phthalate by Fusarium culmorum.

Di(2-ethylhexyl) phthalate (DEHP) is a widely used plasticizer, which is considered an endocrine disrupting pollutant. Growth kinetics and esterases activity by biochemical tests and polyacrylamide gel electrophoresis were characterized for Fusarium culmorum grown in DEHP-supplemented (1000 mg/L) medium as the only carbon source and in control medium with glucose. Intermediate compounds of biodegraded DEHP were identified by GC-MS. F. culmorum degraded 92% of DEHP within 36 h. DEHP was degraded to butanol, hexanal, catechol and acetic acid. It is suggested that the first two compounds would transform into butanediol and the last two would enter into the Krebs cycle and would be mineralized to CO2 and H2O. DEHP induced eight esterase isoforms, which were different to those constitutive isoforms produced in the control medium. It is suggested that five enzymes (25.7, 29.5, 31.8, 97.6 and 144.5 kDa) detected during the first 36 h be involved in the primary biodegradation of DEHP. The rest of the enzymes (45.9, 66.6 and 202.9 kDa) might be involved in the final steps for DEHP metabolism. F. culmorum has a promising practical application in the treatment of DEHP-contaminated environments because it can secrete specific esterase to breakdown high concentrations of DEHP in a short period of time. This research represents the first approach for the study of esterase involved in the DEHP degradation by fungi using this phthalate as the sole source of carbon and energy.

Plasticizers and BPA Residues in Tunisian and Italian Culinary Herbs and Spices.

In the present study, 18 plasticizers and residues in 10 different Tunisian and Italian culinary herbs and spices (black pepper, mint, caraway, coriander, oregano, rosemary, thyme, fennel, verbena, and laurel) were determined by GC/MS. Di-methylphthalate, di-(2-methylpropyl)adipate, di-n-butyladipate, di-propylphthalate, benzylbenzoate, di-phenylphthalate, and bisphenol A were lower than their LOQ in all 118 samples. Among the Tunisian samples, di-(2-ethylhexyl)phthalate was found in all types of samples and di-butylphthalate in 50% of types; all other phthalates were rarely dectected. Among the adipates, only di-methyladipate was found in 90% of types; di-ethyladipate was seldom found and di-(2-ethylhexyl)adipate only in samples of caraway. Di-(2-ethylhexyl)terephthalate was found in all types of samples; di-(2-ethylhexyl)sebacate was detected rarely but with high values. Among the Sicilian samples, di-ethylphthalate and di-(2-methylpropyl)phthalate were detected only in samples of mint; di-methyladipate, di-butylphthalate, and benzylbutylphthalate were identified in oregano and laurel; di-(2-ethylhexyl)terephthalate and di-(2-ethylhexyl)sebacate only in oregano. In any case, the results suggested that intake of these contaminants through spices and herbs is not a dangerous risk to the consumers. Probably, as already observed for the other food, these contaminants could result from pollution of the environment (air, water, and/or soil) and/or farming methods. PRACTICAL APPLICATION: Plasticizers and BPA in Tunisian and Italian spices were determined by GC/MS. Tunisian spices seem to contain more residues than Italian samples. Intake of these contaminants by spices is not a dangerous risk to the consumers. These pollutants could result from environmental pollution or agricultural practices.

Migration of epoxidized soybean oil from polyvinyl chloride/polyvinylidene chloride food packaging wraps into food simulants.

Epoxidized soybean oil (ESBO) has been used in polyvinyl chloride (PVC)/polyvinylidene chloride (PVDC) food packaging cling film as a plasticizer and stabilizer. The aim of this study was to investigate the migration of ESBO from PVC/PVDC cling film, based on gas chromatography mass spectrometry (GC-MS). The specific migration of ESBO was evaluated using various food simulants (water, 4% acetic acid, 50% ethanol and n-heptane) for PVC and PVDC wrap products. ESBO did not migrate into water and 4% acetic acid for all the tested samples. However, it was released into 50% ethanol and n-heptane in several PVC/PVDC wraps, with maximum migration levels of 38.4 ± 0.7 and 37.4 ± 0.8 µg/mL, respectively. These results demonstrate that ESBO is capable of being released from PVC/PVDC wrap into amphiphilic/oily food and its migration should be regularly monitored.

Development of pectin films with pomegranate juice and citric acid.

The influence of pomegranate juice (PJ, replacing water as solvent) and citric acid (CA) on properties of pectin films was studied. PJ provided the films with a bright red color, and acted as a plasticizer. Increasing PJ/water ratio from 0/100 to 100/0 resulted in enhanced elongation (from 2% to 20%), decreased strength (from 10 to <2 MPa) and modulus (from 93 to <10 MPa), increased water vapor permeability (WVP, from 3 to 9 g.mm.kPa(-1).h(-1).m(-2)), and decreased insoluble matter (IM, from 35% to 24%). Although a crosslinking effect by CA was not confirmed, it has been suggested to occur from its effects on films. CA noticeably increased IM (from <10% to almost 40%); moreover, when measured on a dry film basis, the CA effects presented a noticeable tendency to increases strength and modulus, and to decrease WVP. The red color density was decreased by CA, suggesting a destabilization of anthocyanins.

Determination of phthalate esters in vegetable oils using direct immersion solid-phase microextraction and fast gas chromatography coupled with triple quadrupole mass spectrometry.

Phthalates are a group of synthetic compounds mainly used as plasticizers, which have been classified as endocrine-disrupting chemicals and potential human-cancer causing agents. They can be found in high amounts in foods, deriving mainly from plastic packaging. The analytical determination of these compounds is very challenging since they are ubiquitous. Therefore, minimization of sample manipulation is highly desirable. The present work exploited the application of a solid-phase microextraction method for the analysis of phthalates in vegetable oil. A preliminary comparison between a polydimethylsiloxane (PDMS) and a Carbopack Z/PDMS fiber was carried out both in the headspace and direct immersion extraction modes. Before immersing the fiber, a rapid liquid-liquid extraction was performed using acetonitrile to remove the bulk of triglycerides. PDMS in the direct immersion mode showed the best performance. The method was fully validated obtaining a good linearity with a coefficient of correlation of over 0.9960 for all compounds, repeatability and accuracy values generally better than 10%, and very good limit of quantification values.

Migrated phthalate levels into edible oils.

The determination of phthalates in edible oils (virgin olive oil, olive oil, canola oil, hazelnut oil, sunflower oil, corn oil) sold in Turkish markets was carried out using gas chromatography-mass spectrometry. Mean phthalate concentrations were between 0.102 and 3.863 mg L(-1) in virgin olive oil; 0.172 and 6.486 mg L(-1) in olive oil; 0.501 and 3.651 mg L(-1) in hazelnut oil; 0.457 and 3.415 mg L(-1) in canola oil; 2.227 and 6.673 mg L(-1) in sunflower oil; and 1.585 and 6.248 mg L(-1) in corn oil. Furthermore, the influence of the types of oil and container to the phthalate migration was investigated. The highest phthalate levels were measured in sunflower oil. The lowest phthalate levels were determined in virgin olive oil and hazelnut oil. The highest phthalate levels were determined in oil samples contained in polyethylene terephthalate.

Plasticizer contamination in edible vegetable oil in a U.S. retail market.

With the wide application of plastics, the contamination of plasticizers migrating from plastic materials in the environment is becoming ubiquitous. The presence of phthalates, the major group of plasticizers, in edible items has gained increasingly more concern due to their endocrine disrupting property. In this study, 15 plasticizers in 21 edible vegetable oils purchased from a U.S. retail market were analyzed using gas chromatograph-mass spectrometry. Di(2-ethylhexyl) phthalate (DEHP) and diisobutyl phthalate (DiBP) were detected in all oil samples. Benzylbutyl phthalate (BzBP), dibutyl phthalate (DBP), and diethyl phthalate (DEP) were detected at a rate of 95.2, 90.5, and 90.5%, respectively. The detection rates for all other plasticizers ranged from 0 to 57.1%. The content of total plasticizers in oil samples was determined to be 210-7558 µg/kg, which was comparable to the content range in oil marketed in Italy. Although no significant difference (p = 0.05) in the total content of plasticizer was observed among oil species (soybean, canola, corn, and olive), the wider range and higher average of total content of plasticizers in olive oil than other oil species indicated the inconsistence of plasticizer contamination in olive oil and a possible priority for quality monitoring. No significant difference (p = 0.05) in the total content of plasticizers was found among glass-bottle (n = 4), plastic-bottle (n = 14), and metal-can (n = 3) packaging, implying that oil packaging is not the major cause of plasticizer contamination. The daily intake amount of plasticizers contained in edible oil on this U.S. retail market constituted only a minimum percentage of reference dose established by US EPA, thus no obvious toxicological effect might be caused. However, the fact that DEHP content in two olive oils exceeded relevant special migration limits (SMLs) of Europe and China might need attention.

Testing the influence of various conditions on the migration of epoxidised soybean oil from polyvinylchloride gaskets.

Epoxidised soybean oil (ESBO) is widely used as a plasticiser and stabiliser mainly in food contact materials on the base of polyvinylchloride (PVC), especially in the gaskets of jar lids. PVC gaskets containing 10-37% of ESBO were prepared by the baking of PVC plastisols at various process temperatures (180-240°C) in the laboratory. ESBO migration into olive oil and 3% acetic acid was studied at various temperatures (4°C, 25°C, 40°C and 60°C) during a storage time up to 12 months. ESBO released into food simulants was transmethylated, derivatised and analysed by gas chromatography-mass spectrometry (GC/MS). The effect of food processing, i.e. pasteurisation (80°C and 100°C) and sterilisation (125°C) on ESBO migration was also evaluated. The results were critically assessed with respect to the test conditions of specific migration in accordance with the current European Union legislation (Regulation (EU) No. 10/2011). The levels of ESBO migration found confirmed that the test conditions (i.e. 40°C or 60°C, 10 days) representing contact in the worst foreseeable use scenario seem to be insufficient for the simulation of ESBO migration during long-term storage and thus do not provide satisfactory objective results.

Effect of chitosan nanoparticles and pectin content on mechanical properties and water vapor permeability of banana puree films.

UNLABELLED: Puree prepared from over-ripe peeled bananas was used as raw material for films processing in a laboratory padder. Pectin and glycerol as plasticizer were added in small concentrations and chitosan nanoparticles (88.79 ± 0.42 nm medium size) incorporated at 0.2% (dry weight basis) as reinforcement material. The mechanical properties, water vapor transmission, thermal stability, and scanning electron microscopy of fractured film surfaces were characterized. Both pectin and glycerol demonstrated an important role in promoting elongation and film handability as was expected. The incorporation of nanoparticles promoted noticeable improvement of the mechanical properties and acted in reducing the water vapor permeation rate, by 21% for films processed with pectin and up to 38% for films processed without pectin, when compared to the control (puree films with no pectin and nanoparticles additions). Microscopic observation revealed a denser matrix when nanoparticles are incorporated into the films. PRACTICAL APPLICATION: The development of films from fruit purees head to a new strategy for plastic processing from natural resources. The over-ripe or even waste banana can be adequately prepared for batch films processed with reasonable mechanical and barrier properties, suitable for applications in the food segment. The addition of small fractions of chitosan nanoparticles, form nanocomposites enhancing mechanical and thermal stability broadening potential film applications.

The development of thermal nanoprobe methods as a means of characterizing and mapping plasticizer incorporation into ethylcellulose films.

PURPOSE: The phase composition and distribution of ethylcellulose (EC) films containing varying amounts of the plasticizer fractionated coconut oil (FCO) were studied using a novel combination of thermal and mapping approaches. METHODS: The thermal and thermomechanical properties of films containing up to 30% FCO were characterized using modulated temperature differential scanning calorimetry (MTDSC) and dynamic mechanical analysis (DMA). Film surfaces were mapped using atomic force microscopy (AFM; topographic and pulsed force modes) and the composition of specific regions identified using nanothermal probes. RESULTS: Clear evidence of distinct conjugate phases was obtained for the 20-30% FCO/EC film systems. We suggest a model whereby the composition of the distinct phases may be estimated via consideration of the glass transition temperatures observed using DSC and DMA. By combining pulsed force AFM and nano-thermal analysis we demonstrate that it is possible to map the two separated phases. In particular, the use of thermal probes allowed identification of the distinct regions via localized thermomechanical analysis, whereby nanoscale probe penetration is measured as a function of temperature. CONCLUSION: The study has indicated that by using thermal and imaging techniques in conjunction it is possible to both identify and map distinct regions in binary films.

Detection methods for autologous blood doping.

The use of blood doping is forbidden by the World Anti-Doping Agency. Several practices, such as blood transfusions are used to increase oxygen delivery to muscles and all of them are highly pursued. In this regard, the development of accurate methodologies for detecting these prohibited practices is one of the current aims of the anti-doping control laboratories. Flow cytometry methods are able to detect allogeneic blood transfusions but there is no official methodology available to detect autologous blood transfusions. This paper reviews protocols, including the Athlete Biological Passport, that use indirect markers to detect misuse of blood transfusions, especially autologous blood transfusions. The methods of total haemoglobin mass measurements and the detection of metabolites of blood bags plasticizers in urine are reviewed. The latter seems to be an important step forward because it is a fast screening method and it is based on urine, a fluid widely available for doping control. Other innovative approaches to blood transfusion detection are also mentioned. A combination of the reported methodologies and the implementation of the Athlete Biological Passport is becoming a promising approach.