Plastic, plastic everywhere...but not a drop to drink

Life in today’s modern world would not be possible without plastic, but some of the by-products of its manufacture are far from environmentally friendly. Here we learn how to determine levels of phthalates in drinking water by Auto-SPME and GC-MS.

GLOBAL annual consumption of plastic material has increased by 20 fold from that of 50 years ago, and in the UK alone, a total of nearly 5 million tonnes of plastic products were used at the turn of the 21st Century. Since the first plastics were synthesised in the 1860s, scientists have developed many varieties, and these can be found in a wide range of consumer products, and its versatility allows it to be used in everything from car parts to toys, and from soft drink bottles to the refrigerators they are stored in. However, the polymers that are known as plastics would be a lot harder and more difficult to use if it weren’t for the addition of chemicals known as Phthalic Acid Esters (PAEs), also known as phthalates.

Phthalates are a family of compounds made from alcohols and phthalic anhydride, and are key additives that act as intermolecular lubricants to increase polymer flexibility, keeping the plastics soft at room temperature. They are colourless, odourless liquids that show low water solubility, high oil solubility and low volatility. Due to their widespread use and manufacture in plastics, phthalate esters are one of the most ubiquitous classes of compounds in today’s environment, and can be found as components of cosmetics, detergents, building products, lubricating oils, and carriers in pesticide formulations and solvents. There are many different kinds of phthalates that can be used in plastics and Di (2-ethylhexyl) Phthalate, or DEHP, is the most widespread phthalate produced and used. The greatest use of DEHP is as a plasticiser for polyvinylchloride (PVC) and other polymers including rubber, cellulose and styrene. DEHP is also primarily used in the production of a number of packaging materials and tubings for foods and beverages.

Nowadays, phthalates are considered as environmental pollutants. Since phthalates are physically rather than chemical incorporated into the polymeric matrix structure, they are highly mobile and can separate from the plastic products. Thus, relatively large amounts of these compounds can easily migrate from the packaging or manufacturing processes and released into the environment, eventually making their way into food, drinking water and the air. Although the highest levels tend to be found in fatty foods and vegetable oils, the presence of phthalates in surface, ground and drinking water can occur at trace levels, which is linked to contamination release from chemical plants. Consequently, the potential for human exposure is very high and in recent years, considerable attention has been paid to phthalates because of their suspected carcinogenic and estrogenic properties. Toxicological studies have also linked some of these compounds to liver and kidney damage, as well as to possible testicular or reproductive-tract birth defect problems, characterising them as endocrine disruptors.

Although scientists have long understood that bodies absorb tiny amounts of chemical substances simply by interacting with the environment, today’s technology allows researchers to detect and measure trace concentrations of many environmental substances in the body. Scientists have now developed very sensitive tests that can find minute traces - a millionth of a gram or even less - of certain chemicals or their metabolites. Relatively large amounts of phthalates are released to the environment, and because some phthalates leach more readily than others and are found in higher concentrations in air, food and water, the identification of specific phthalates is important and a better knowledge of their biological impact is needed. Therefore, in order to study this impact, reliable quantification methods are required.

There have been many sample preparation methods that have been purposed for the determination of phthalates in water samples for enrichment prior to detection with techniques such as gas chromatography (GC), mass spectrometry (MS) or high liquid performance chromatography (HPLC). These extraction methods, including liquid-liquid extraction (LLE) and solid-phase extraction (SPE), utilise solvents in the sample preparation step, such as hexane and dichloromethane, and have been widely used to isolate phthalates from aqueous samples. However, these procedures are typically time-consuming, labour-intensive, and use a large amount of solvent. Moreover, from a production point of view, industries using phthalates need a more complete characterisation of their starting materials and final products to maintain appropriate phthalate compositions and concentrations. Unfortunately, this is very difficult using traditional chemical analysis.

Therefore, in the last decade, solid-phase microextraction (SPME) has become a popular method for sample preparation and has been successfully applied for the analysis of phthalates in water matrices. SPME is a fast, sensitive, solventless, and economical sample preparation method for gas chromatography analysis. The main advantages of SPME compared to solvent extraction are the reduction in solvent use, the combination of extraction and analysis into one step, and the ability to examine smaller sample sizes. It can also provide high sensitivity and can be used for polar and non-polar analytes in a wide range of matrices with direct injection to both the gas chromatograph (GC) and the liquid chromatography (LC).

Agilent have devised an approach to determine plasticisers in water samples, and an automated SPME sample preparation process is demonstrated by using an Agilent CombiPAL combined with GC-MS. Extraction of analytes from aqueous samples can be performed either by direct immersion of the fibre into the liquid phase or by headspace sampling. Adsorbed analytes are then thermally desorbed in the injection port of a GC and analysed using an appropriate column and detector. All movements of the SPME fibre from precondition, adsorption, and desorption are software controlled for optimum precision. Prior and during extraction, the samples can be shaken and heated. This approach dramatically reduces sample preparation time for semi-volatile compounds. Variable vial penetration depth allows compound extraction to be performed in liquid phase or in the headspace. After the compounds are thermally desorbed in the hot GC injector, the fibre may be regenerated in a heated and purged cleaning station.

The experiment
A solid-phase microextraction (SPME) method for the analysis of phthalates in drinking water samples has been developed on the Agilent CTC CombiPAL autosampler GC-MS platform. In this method, the sample preparation process was automated by using a CombiPAL autosampler, including the SPME fibre precondition, adsorption, and desorption, which improve the precision of the SPME method. The extraction temperature, extraction time, and salt-out effect are also studied. The optimized condition was applied to the analysis of real samples. The detection limits of the phthalates in this method are at the sub-ppb level.  The RSDs for most of the esters are less than 10%, the detection limits calculated by 3 times of S/N are less than 0.5ng/ml.

CombiPAL
Pre-incubation time: 60 s
Incubation temperature: 40 °C
Pre-inc. agitator speed: 500 rpm
Agitator on time: 5 s
Agitator off time: 2 s
Vial penetration: 25 mm
Extraction time: 1200 s
Desorb to: GC Inj1
Injection penetration: 54 mm
Desorption time: 120 s
Post fiber condition time: 300 s

SPME
SPME fiber is from Supelco company (595 North Harrison Road Bellefonte, PA, USA), the fiber type is polydimethylsiloxane/divinylbenzene (PDMS/DVB) and the coating thickness is 65 µm.

6890 GC
Inlet temperature: 270 °C
Gas type: Helium
Oven condition: 50 °C Ramps 10.00 °C /min to 260 °C (3.00 min)
Column: DB-5ms 30 m × 250 mm, 0.25 µm
Mode: Constant flow
Flow rate: 1.3 mL/min

5975 MS
Acquisition mode: Synchronous SIM/scan
Mass range: 40–300
Sample: 3
Dwell time: 30 ms
MS source: 230 °C
MS quad: 150 °C
For other parameters, see Table 1.

The PAEs standards (shown in Table 1) were bought from Guo Yao Group (Shanghai, China). 

Results and Discussion
Because PAEs are semi-volatile compounds, immersion extraction mode was selected, and the sample volume was 18mL.
Lots of unrelated peaks emerged in GC chromatograms when the extraction temperature was over 40°C, which would shorten the lifetime of fibre, so a compromise has to be made between the lifetime of the extraction phase and the rate of equilibrium. We chose 40°C for all extractions in the following experiments.

The effect of extraction time versus amount extracted at 40°C was studied. The extraction efficiency for different compounds was proportional to extraction time. Figure 1 shows the profile of extraction time versus response. As seen in Figure 1, when the extraction time was over 20 minutes, the responses changed slightly, which means that the extraction of most compounds reached equilibrium at this point. In this experiment, 20 minutes was selected as the extraction time.

Salting-out effects by adding NaCl in the sample were also studied. The results showed that the extraction efficiency of DEP, DMP, and DBP was improved when salt was added, and that of DCHP, DEHP, and DPP (see compound names in Table 1) was decreased as shown in Figure 2. In this experiment, 20% (W/V) salt concentration was chosen. Figure 3 shows the SIM chromatogram of PAEs at the optimised condition. The chromatogram shows that improvements can be made to shorten the analysis time by adjusting the oven program.

The linearity of the analytes was determined by calibration solutions with the concentration range from 0.5 ppb to 1 ppm at the optimized extraction condition. Table 2 shows the concentration ranges and correlating coefficients. The precision of the analysis, represented as relative standard deviations (RSDs) at 1 ppb, is also shown in Table 2. The RSDs for the organic esters are less than 10% except that of DPP; the detection limit is calculated at S/N of 3.
To demonstrate the performance of the optimized SPME method, tap water, potable water, and purified water from a water dispenser were analysed for the phthalates’ presence. Table 3 shows the phthalates detected in these three samples.

Conclusions
The CombiPAL autosampler with SPME is used for the analysis of PAEs in water. The precondition, extraction, adsorption, and desorption of SPME are automated and precisely controlled, which improves the precision of SPME method. Because the analytes concentrate into the coating of SPME, trace-level contaminates can be detected by using SPME. In this application, the detection limits for PAEs are down to sub-ppb level.

The United States Environmental Protection Agency (USEPA) under the Safe Drinking Water Act regulates DEHP contaminant levels at 6.0µg/L, and this method could meet the demands of EPA’s regulation from the detection limits perspective. This developed solution is an effective and environmentally friendly method for the detection of PAEs in water samples and also has the potential to be used in other samples.