Leaching of Phthalate from Medical Devices use in Pakistani Hospitals

Phthalates (PAEs) are widely used as plasticizers in medical devices to make polyvinyl chloride flexible and soft. However, PAEs can be leached out from plasticizers undesirably and can migrate within the material over the time and can end up with direct exposure to humans, because PAEs are not chemically bound, Particularly, (2-ethylhexyl) phthalate (DEHP) is present within various industrial and domestic goods along with medical devices. Leaching of PAEs is the main concern for human health due to toxic health effects, such as endocrine disruptor (EDCs) carcinogenicity, breast cancer, asthma, allergies and so on. PAEs are produced all over the world around 6.0 million metric ton per PAEs expose specifically from plastic containers, like cooler, plastic bottles, soft drinks, mineral water bottles and baby feeders and medical devices. study the leaching of PAEs from medical devices abundantly use in local hospitals. After market survey in first step, infusion (IF) bottles of various brands available for infusion of dextrose, normal saline and mannitol packed in plastic bags/ packing along with this catheters and syringes were collected and coded to fulfill ethical requirements. Total 30 samples (103) of IF, 9 (3x3) samples of catheters along with 3 samples of syringes were collected and analyzed. The IF samples were classified on the basis of their composition, such as sugar (glucose/mannitol) and salt. Leached PAEs were extracted with dichloromethane following (EPA) standard method (606). Moreover DSC analysis showed clear Tg and Tm values for the infusion bottles, which lies in the category of amorphous nature. In addition IF bottles catheters and syringes were also analyzed by FTIR characterization. Results indicated that all IF solutions were acidic with pH ranges from 2.34 to 5.88. Gas chromatographic analysis showed that extent of leaching was different among brands; normal saline showed more PAEs leaching (78.60 µg/L) followed by dextrose (34.29 µg/L), mannitol (20.93 µg/L). In addition leaching studies of catheters was carried out with synthetic saline solution; results showed the total PAEs leached were in the range of 3.96 to 51.484 µg/L. In syringes PAEs leaching was not detected by Gas chromatography. catheters additional 2 PAES i.e DiBP and DiProP. The Overall study indicated that due to medical devices such as infusion solutions, human are at direct exposure risk of phthalate accumulation. Dipropyl Phthalate; Dibutyl Phthalate; Phthalate; BBP: Benzylbutyl Phthalate; Phthalate; Phthalate;

medical gloves [2]. PAEs are abundantly used in many goods like paints, fragrances, nail polish, skin care products, food packing, hair products cosmetics and printing inks [3]. The general figure of PAE is depicted in 1.1. In MDs Phthalates (PAEs) like di-(2-ethylhexyl) (DEHP) is used as a major source which is about 30 to 40% of its weight [4]. In Europe about one million tons of PAEs are produced per year and the dominant PAEs are diisononyl phthalate (DiNP), diisodecyl phthalate (DiDP) and di-(2-ethylhexyl) phthalate (DEHP) are used as plasticizers [5]. PAEs are not chemically bound to plastic therefore leached into plastic material and spread into environment frequently [6]. PAEs are released out from MDs quickly by high temperature, lower pH, duration of use, infusion rate, and surface contact [7]. Human beings are highly affected by the PAEs over the larger scale [8] [DEHP, benzyl-butylphthalate (BBP) and dibutyl-phthalate (DBP)] have mutagenic and carcinogenic impact over the breastfeeding or pregnant women and children [9]. The DEHP, benzyl butyl phthalate (BBP) and dibutyl phthalate (DBP) are known as carcinogenic endocrine disruptors to humans and some of their studied showed their effects of mutagenic activity, peroxisome proliferation, infertility and carcinogenicity [10]. There are numerous ways through which human beings are exposed to PAEs such as foodstuff, dermal resorption or by inhaling air which contains PAEs [11].
The leaching and toxicity studies of PAEs were carried out by Duty et al [12], Schug et al; [13]; they reported the great impact of PAEs over the health of human beings through their studies.
Previously the leaching study of PAEs from the PVC bags was reported by Bernard et al; [14], infusion sets Inoue et al [15], PVC tubing Takehisa et al; [16], blood bag shaz et al [17]. The Toxic and neurological effect over the animals and human beings [18] even can affect their liver [19]. The children are highly affected by the PAEs which results the great damage in their brain function. Moreover, the sperms of human beings along with their DNA are damaged due to the diethyl hexyl phthalate (DEHP) and DEP. The PAEs results in the destruction of lungs reported by Schutze et al; [13]. Along with this the exposure to PAEs from PVC material causes the enlargement of allergies, asthma [20]. The reproduction effects showed that the DEHP reduces the testicular weight as well as production of sperms and contributes to the atrophy of seminiferous tubules which results to infertility and damage kidney liver, spleen cancer and thyroid in male rodents [21] and PAEs lead to the disorders in the development of reproduction and in testicular dysgenesis in humans [22], keeping in view about the health effects of PAEs many of the countries like France in 2015 banned tubes used in neonatal possessing DEHP [23], US in 2012 [24], Canada in 2010 [25], USA in 2008 [26] and European union in 2005 [27] has banned PAEs to be used in PVC as plasticizers. Currently many of the countries like United States, Germany, Switzerland and Austria use the alternative of PAEs like Diisononyl cyclohexane dicarboxylate (DINCH) has substituted the DEHP [13]. The common alternates of PAEs are excessively used as free plasticizers in medical devices which have no health effect such as Diisononyl adipate (DINA), Bis(2ethylhexyl) terephthalate (DEHT) [28,29].
In Pakistan generally the temperature reaches up to 50oC and it is reported in literature with the increasing effect of temperature the leaching from the PVC bottles takes place at higher rate [30].
Since MDs have direct impact on human health in our country due to having high temperature and MDs are stored at room temperature. The pH has direct impact over the leaching of PAEs from the MDs [31] and all the IF bottles are found to be in lower pH.

Extraction of Leached Phthalates
Leaching of PAEs from the MDs was followed by liquid -liquid extraction as per reported method presented by Suhrhoff [32].
The 0.9% solution of normal saline was prepared and passed through catheters and syringes. 10 ml of each infusion solution was extracted with 20 ml DCM and extraction was repeated thrice for maximum recovery of leached PAEs. The anhydrous sodium sulphate (Na 2 SO 4 ) was used to dry the combined organic phases.
The gas chromatography (GC) combined along with the flame ionization detector (FID) was used to analyze samples.

Gc -Fid Conditions for the Determination of Paes.
The leached PAEs were monitored by gas chromatograph The DB-1 (30m x 0.25 mm ID) m long polar capillary column, with the gas chromatograph (GC-8700 PERKIN ELMER) was used to analyze the substrate and as a carrier gas nitrogen was utilized, temperature of oven was raised from 120 to 220oC with the ramp rate of 4oC /min and final temperature 220oC was set for 30 minutes, 2 µL volume was used as injection. The temperature of detector and injector was set at 240°C and 260°.

Identification of Leached Paes By Gc -Ms
The leached PAEs from MDs were identified by using Agilent

Mds Characterization by Differential Scanning Calorimetry (Dsc)
The thermal behavior of PVC was analyzed by using differential scanning calorimetry (DSC). The rate of heating was 7°C/min in the range 60 to 150°C was set by using the stream of N 2 gas (10 mL/min). In sample pan approximately the 5 mg of PVC tubes were sealed. The midpoint of a small endothermic rise of the pre-and post-transition baselines was used to determine the glass transition temperature (Tg) of PVC films, method described by Takehisa [16].

Ftir -Atr Characterization of Plastic Bottles
All the plastic infusion bottles syringes and catheters were studied by FTIR -ATR analysis in the region of 400 -4000 cm -1 .     (Table 3.2).

Gc -Ms. Quantification of Leached Phthalates
The identification of PAEs was also carried out by the GC -MS by measuring the mass fragments and molecular ion peak. The general  [36] for detection of phthalates in bottled water by gas chromatography-mass spectrometry [37]. All the mass fragments of PAEs which were leached are present in (Table 3.3) and the mass spectra of leached PAEs are depicted in (Figure 3.3a and 3.3 b).

Mds Characterization by Differential Scanning Calorimetry (Dsc)
DSC analysis measures the heat absorbed when the temperature of the sample is raised at the linear rate and the process can be either exothermic or endothermic along with its morphology [38].
The experiment provides the (Tg) glass transition temperature and the percent crystallinities to assess the morphological state i.e (amorphous, crystalline and oriented). At the (Tg) the polymer is in its glassy state at which cold draw and plastic deformations are possible as the temperature is raised the secondary interaction between the adjacent polymers become weaker and the polymer lost its glassy state and is completely amorphous state [39].  [16] which proved the leaching can take place at this range and this is due to the amorphous nature of samples.
The DSC thermogram of representative samples i.e. normal saline, mannitol, sugar, syringe and catheter ( Figure 3.4 a,b c,d and e) respectively are given with details with transition and melting temperatures in (Table 3.4).

Conclusion
The present study has revealed that phthalates leaching from infusion bottles and catheters syringes were found at different ranges, while no PAEs leaching was found from different syringe

Competing Interests
All the authors declare that they have competing interest.