Closing in on DEHP‐free red blood cell concentrate containers

For almost 50 years,1 transfusion science has been in search of a replacement for the di-2-ethylhexyl phthalate (DEHP) plasticizer that is used in our polyvinyl chloride (PVC) whole blood collection and red blood cell (RBC) storage containers. The introduction of plasticized PVC containers in the 1950s improved the safety of blood for transfusion by decreasing bacterial contamination and preventing container breakages, reducing hemolysis, and improving in vivo cell recoveries.2, 3 As a durable but flexible material that can be steam sterilized and heat welded and is strong enough to tolerate centrifugation and compressive pressures, DEHP PVC plastic containers quickly replaced glass bottles. This advanced blood banking efficiency and ultimately paved the way for the development of blood component manufacturing.4 While PVC DEHP remains the most popular material used in RBC storage containers,5 the toxicity of DEHP and its congener phthalate metabolites has been and remains a topic of public concern.6, 7 DEHP, a known toxin and endocrine-disrupting compound,7 is used widely in PVC-based medical devices, toys, food packaging, and other consumer goods. DEHP does not chemically bond to the PVC plastic and therefore will leach into the blood components, where it can be infused into the blood donor during apheresis collections or into the patient during a transfusion. Concerns over the toxicity of DEHP and its metabolite mono ethylhexyl phthalate (MEHP), and their association with long-term adverse reproductive, cardiac, metabolic, and neurologic health effects, has been well documented.7-9 The potential for these phthalates to induce reproductive toxicity, infertility, and carcinogenicity is supported by in vitro and in vivo animal studies.6, 7, 10 As specific patient groups, such as neonates, pregnant women, children, and adult patients, who require massive or frequent transfusions, are considered at a greater risk of DEHP exposure, a case for taking active precautionary measures to reduce or eliminate exposure has been made.6, 7, 11 Given these concerns, regulatory agencies and governments have been active in evaluating the risks associated with phthalate exposure and have enacted legislation to more tightly regulate its use in consumer and medical products.7, 12 Concerns over potential long-term effects of DEHP in blood containers have instigated the search for alternative plasticizers. For RBC components, replacing DEHP by a nonphthalate PVC plasticizer is challenged by the membrane protective effect that DEHP exerts on RBCs ex vivo.13 DEHP has been shown to improve stored RBC morphology,14 deformability,15 osmotic fragility,2, 15 and microvesicle release16 without affecting 2,3-diphosphoglycerate or adenosine triphosphate levels.14 The enhanced RBC in vivo recovery observed with DEHP3 underlines the plasticizer's role in preserving the plasma membrane during storage. Although DEHP-free, PVC-free plastic bags with improved gas diffusion characteristics have been introduced for platelets, and plastic bags with better breakage resistance have been developed for frozen plasma storage, the options available for RBC containers have been limited.5 Alternative plasticizers for use in RBC storage containers have included BTHC [butyry-trihexyl-citrate], DINCH [di(isononyl) cyclohexane-1,2-dicarboxylate], and DEHT [di-(2-ethylhexyl) terephthalate].17-19 While the citrate-based BTHC has been used extensively in platelet storage containers due to its excellent gas permeability characteristics, it is available in only a limited number of commercial RBC containers.5 BTHC leaches more slowly from PVC plastic, has a low toxicity, and has metabolites (hexanol, butyric acid, and citric acid) that are unlikely to accumulate in the body. However, several drawbacks exist for this plasticizer compared to DEHP, including a lower protective effect for RBCs (higher hemolysis) requiring a lower storage duration (35 days),17, 20 higher price, unpleasant odor, and an inability to be steam sterilized.21 On the other hand, DINCH has very similar structural and technical properties as DEHP without the negative toxicity effects.21 DINCH has been shown to be a better alternative than BTHC plasticizer because it retains the same RBC membrane–stabilizing effects as DEHP and can maintain RBC quality for up to 42 days of storage.13, 20 Therefore, although DINCH is more expensive than DEHP, due to its favorable toxicity profile, it is considered as one of the most promising alternative plasticizers to DEHP.13 In this issue of TRANSFUSION, Graminske and colleagues18 report on the use of DEHT as a replacement for DEHP in RBC storage containers. Unlike DEHP, DEHT undergoes complete hydrolysis in vivo, which affords it an improved toxicological profile with no reported carcinogenic, mutagenic, or reproductive effects.18, 22 Using a pool and split study design, Graminske and colleagues18 show that storage of RBCs in DEHT-PVC/PAGGSM (phosphate-adenine-glucose-guanosin-saline-mannitol) has comparable levels of hemolysis, supernatant potassium, and RBC biochemical properties after 42 days of storage as RBCs stored in DEHP-PVC/AS-1 containers. Differences in RBC morphology, mean cell volume, pO2, and glucose consumption were noted between the two groups, but these differences were deemed not to be clinically relevant. Interestingly, the study demonstrates that storage for 42 days in DEHT-PVC/AS-1 containers did not afford the same storage stability as the DEHT-PVC/PAGGSM, which highlights the synergistic role that exists between the choice of plasticizer and additive solution.23 While further in vitro studies with different additive solutions and in vivo circulation studies need to be performed to establish the safety and efficacy of RBC concentrates stored in DEHT-PVC containers, the results of this study would suggest that DEHT-PVC is a viable replacement for DEHP-PVC. The risks associated with the use of DEHP-PVC blood storage containers have been widely debated due to the limited human data to directly support a link among the reproductive, cardiac, metabolic, and neurologic health effects that have been associated with exposure to phthalates. However, as Shaz and colleagues6 and others7, 11 have argued, the precautionary principle would demand that any potential risk to blood donors and recipients requires transfusion services to explore and adopt solutions that minimize or reduce the risk as soon as they are available. Efforts to reduce the use of DEHP-PVC plastics in platelet, plasma, and hematopoietic stem cell storage containers have been widely implemented.5 Unfortunately, as highlighted in recent surveys of blood manufacturers,11, 24 the lack of availability of viable alternatives to DEHP-PVC containers for the storage of whole blood or RBC concentrates has prevented their widespread adoption. The unanticipated success4 of DEHP-PVC plastics at preserving RBC in vitro and in vivo quality during extended storage has created the current challenge we have with identifying alternative plasticizers. Nontoxic alternatives to DEHP-PVC must meet or exceed the current performance characteristics that regulators, blood operators, and clinicians expect with minimal change in their practices. While promising results have been demonstrated with DINCH, and now DEHT plasticizers, it is clear that these alternatives do not achieve the same in vitro and in vivo storage characteristics as DEHP when compared directly with existing products in the market. The solution to removing DEHP from whole blood and RBC storage bags may need to involve not just the identification of alternative plasticizers but also parallel adoption of new additive solutions, the introduction of alternate manufacturing methods that are less stressful to RBCs, and/or a shortening of the RBC storage period. In the absence of DEHP to stabilize the RBC membrane, it has been suggested that next-generation additive solutions might be able to compensate for this lack of stabilization when alternate plasticizers are used.23 It has been shown that the use of PAGGSM, AS-3, or PAGGGM in DINCH-23 or DEHT-plasticized18 collection and storage systems perform similarly to DEHP-PVC containers with AS-1 or SAGM (saline-adenine-glucose-mannitol). It is therefore prudent that exploration of the synergistic effect of next-generation additive solutions, such as AS-7, E-Sol, or PAGGGM, with DEHP-free container systems is undertaken. The storage characteristics of RBCs, including hemolysis, deformability, microvesiculation, and morphology, can be significantly affected by the choice of apheresis or whole blood manufacturing method used by a blood service.25-27 Storage hemolysis in products manufactured using RBC filtration (top/bottom, or buffy coat) can be significantly better than whole blood filtered (top/top) or apheresis RBC concentrates.26 When evaluating alternatives to DEHP, it may be valuable to consider the impact that blood component manufacturing may be contributing to the RBC stability characteristics. While current alternative plasticizers (DINCH, DEHT) have not shown equivalent levels of end-storage hemolysis as DEHP-PVC containers, the levels of hemolysis and RBC storage injury at earlier storage periods (28 or 35 days) are comparable. If initiatives28 to reduce the current storage period for RBCs to 35 days are successful at changing practice, then the adoption of existing DEHP-free container systems could be considered. A recent survey24 of scientific professionals working in the field of blood transfusion suggests that respondents would compromise on either storage time or in vitro quality (i.e., higher hemolysis) if regulatory agencies required a switch to non-DEHP plastics. However, the current increased cost of DEHP-free container systems was seen as a barrier unless they became mandated by regulatory agencies or legislation. If a change in additive solution or manufacturing method were required to meet quality standards, then support from industry and the regulator would need to be provided to allow their adoption at the same time as the implementation of DEHP-free containers. Alternatives to DEHP plasticizers with reduced toxicity and comparable RBC stabilizing effects are available. The study by Graminske and colleagues18 highlights the significant advances that have been made using alternative plasticizer and additive solutions that can provide equivalent maintenance of RBC quality. However, adoption of these whole blood or RBC storage containers may require blood systems to adopt additional changes in their operations to fully achieve the in vitro and in vivo quality characteristics of existing DEHP-PVC containers. Because the argument that there are no suitable replacements to DEHP would appear to be no longer true, it may only be a matter of time before we begin to realize the evasive 50-year goal of eliminating DEHP from blood bags is ultimately achievable. The authors have disclosed no conflicts of interest. Ruqayyah J. Almizraq, BSc, MSc1 e-mail: ruqayyah@ualberta.ca Jason P. Acker, BSc, MSc, MBA, PhD1,2 e-mail: jason.acker@ualberta.ca 1Department of Laboratory Medicine and Pathology University of Alberta, Edmonton, AB, Canada 2Centre for Innovation, Canadian Blood Services Edmonton, AB, Canada

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