G. Bersani

Vitamin A and vitamin E isoforms stability and peroxidation potential of all-in-one admixtures for parenteral nutrition.

BACKGROUND In all-in-one admixtures (AIOs), vitamins can be degraded and lipid can be peroxidized by light exposure, oxygen action, and multiple chemical interactions. AIM We investigated the impact of three commercial lipid emulsions and two multivitamin preparations on vitamin A and vitamin E chemical stability and lipid peroxidation potential of AIOs. METHODS A soybean oil (Soy), soybean/medium-chain triacylglycerol oil (MCT), and olive/soybean oil (Olive)-based emulsion (all 20%), and a lyophilized (Lyo) and emulsified (Emu) multivitamin compounds, were tested. Two AIOs for each lipid emulsion were prepared, the former with Lyo and the latter with Emu. The concentrations of retinol palmitate, alpha-gamma-delta-tocopherol, and malondialdehyde were analyzed in AIOs, immediately (T0) and 24 hours (T24) after compounding. RESULTS Retinol palmitate, and alpha- and gamma-tocopherol were more stable in MCT-AIOs than in both Soy-AIOs and Olive-AIOs (p < 0.013; p < 0.001 respectively). Furthermore alpha-tocopherol was more stable in Lyo-AIOs than in Emu-AIOs (p < 0.004). Malondialdehyde (MDA) increased differently among the admixtures; however the concentrations were similar in all AIOs at T24. CONCLUSIONS The differences in retinol palmitate stability were due both to lipid emulsions per se and to interaction between lipid emulsions and multivitamin preparations. The alpha-gamma-tocopherol stability depended on both lipid emulsions and multivitamin preparations. In tested AIOs there was a different degradation rate of fat-soluble vitamins to keep the same lipid peroxidation level, since MDA concentrations at T24 were similar among AIOs.

Phytonadione Content in Branded Intravenous Fat Emulsions A Comparative Study of 6 Products With Different Fat Sources Using Liquid Chromatography–Mass Spectrometry

Background: Intravenous fat emulsions (IVFE) with different fatty acid compositions contain vitamin E as a by-product of vegetable and animal oil during the refining processes. Likewise, other lipid-soluble vitamins may be present in IVFE. No data, however, exist about phytonadione (vitamin K1) concentration in IVFE information leaflets. Therefore, our aim was to evaluate the phytonadione content in different IVFE. Materials and Methods: Analyses were carried out in triplicate on 6 branded IVFE as follows: 30% soybean oil (100%), 20% olive-soybean oil (80%–20%), 20% soybean–medium-chain triglycerides (MCT) coconut oil (50%–50%), 20% soybean-olive-MCT-fish oil (30%-25%-30%-15%), 20% soybean-MCT-fish oil (40%-50%-10%), and 10% pure fish oil (100%). Phytonadione was analyzed and quantified by a quali-quantitative liquid chromatography–mass spectrometry (LC-MS) method after its extraction from the IVFE by an isopropyl alcohol–hexane mixture, reverse phase–liquid chromatography, and specific multiple-reaction monitoring for phytonadione and vitamin d3 (as internal standard). This method was validated through specificity, linearity, and accuracy. Results: Average vitamin K1 content was 500, 100, 90, 100, 95, and 70 µg/L in soybean oil, olive-soybean oil, soybean-MCT coconut oil, soybean-olive-MCT-fish oil, soybean-MCT-fish oil, and pure fish oil intravenous lipid emulsions (ILEs), respectively. The analytical LC-MS method was extremely effective in terms of specificity, linearity (r = 0.99), and accuracy (coefficient of variation <5%). Conclusions: Phytonadione is present in IVFE, and its intake varies according to IVFE type and the volume administered. It can contribute to daily requirements and become clinically relevant when simultaneously infused with multivitamins during long-term parenteral nutrition. LC-MS seems adequate in assessing vitamin K1 intake in IVFE.

Phytosterols and Lack of Occurrence of Cholestasis in Rats Nourished Parenterally or Orally

Intravenous lipid emulsions (ILEs) have been marketed to infuse fat within total parenteral nutrition (TPN) in order to prevent essential fatty acid deficiency. Only recently, ILEs have been recognized to have therapeutic effects in gastrointestinal, cardiovascular, pulmonary, oncologic, autoimmune, and critical care diseases. At the same time, toxic substances such as phytosterols have been found in ILEs. Phytosterols have been related to a TPN-associated complication defined parenteral nutrition-associated cholestasis (PNAC) (1-5).

Insulin Instability in Parenteral Nutrition Admixtures

BACKGROUND Biosynthetic human recombinant short-acting insulin is added to parenteral nutrition (PN) admixtures to nourish glucose-intolerant patients. Insulin, however, is electrostatically attracted and inactivated by ethyl-vinyl-acetate (EVA) bags and filling system tubes. Our aim was to verify and quantify the presence of insulin in PN with and without intravenous lipid emulsion (ILE), just after addition (T0) until the infusions end (T24). METHODS Four undiluted samples of 12 different PN complete admixtures (6 with ILE and 6 without), each containing 250 g of glucose in a 2000 mL volume, were taken and analyzed at T0 and T24 by an automated electrochemiluminescence immunoassay after the addition of biosynthetic human recombinant short-acting insulin at increasing doses (from 6 to 72 IU/bag) by an automated compounding device. Assay sensitivity was set at 2 μIU/mL. Admixtures with and without ILE were compared in terms of insulin-detected amounts at T0 and T24. RESULTS Regardless of the amount initially provided, insulin was missing in PN without ILE. In admixtures with ILE, the greater the insulin and ILE doses initially included, the higher the insulin availability at T0 and T24, both in absolute terms and as a percentage of the initial amount (from 3 to 81% at T0 and from 2.5 to 72.5% at T24). ILE may prevent insulin attraction to plastic surfaces. CONCLUSIONS Insulin is recovered in the presence of ILE in PN even though considerable amounts are untraceable. This aspect needs verification. Until then, insulin should safely be injected in a different manner in uncontrolled situations.

PP-036 Preparation of an experimental cord blood serum eye drops for topical use in severe corneal epitheliopathy

Background The use of cord blood (CB) serum as a tear substitute has been recently proposed to heal severe corneal epithelial damage due to its high concentration of epithelial growth factors. No branded CB serum-based eye drops are available on the market. Purpose To optimise the preparation of experimental galenical CB serum-based eye drops by ensuring product safety. Material and methods In S. Orsola-Malpighi Hospital of Bologna, Italy, the drops were made under a vertical laminar flow hood. In the Transfusion Centre serum was collected from CB units, centrifuged for 10 min at 3,500 rpm, aliquoted into 15 ml sterile tubes and frozen at -80°C for six months as a quarantine period. Serological and molecular tests were performed on each serum sample, according to the Italian regulations. Thawed CB serum was sent to the pharmacy laboratory where it was diluted 1:5 with refrigerated sterile physiological saline, filtered (Millex HV 0.45 μm) and aliquoted into 1 ml single-dose PET vials. Filled vials were thermo-welded and packed in sealed labelled envelopes. Finally, they were stored at −20°C for 30 days before delivery to patients. Certification of molecular and serological tests was retained in the pharmacy. CB serum levels of epithelial growth factors were tested at different steps: freshly collected, thawed after quarantine, after filtration, after dilution and after one or two months storage at −20°C, respectively. Sterility was validated by a BacT/Alert test on each batch of eye drops. Results Sterility tests confirmed that all batches of eye drops remained sterile after handling and storage. Immunological tests showed that CB serum levels were maintained over the whole process. Patients must keep the eye drops refrigerated and use them within 12 days. Conclusion The collaboration among interdisciplinary professional figures overcame critical preparation problems, providing patients with a safe and effective product. References and/or acknowledgements No conflict of interest.

Replay to the Editor regarding Prof Klek's letter -reference manuscript # YCLNU-D-13-00298R2 Parenteral Nutrition Admixtures for Pediatric Patients Compounded with Highly Refined Fish Oil-Based Emulsion: Assessment of Physicochemical Stability.

I am pleased to respond to Prof. Klek regarding his comments on our article. His substantial concerns were related to the safety limit of 4.5 mmol/L of calcium concentration found in parenteral nutrition compounded with different lipid formulations and the analytical method we used. First of all, we think we have pointed out in the manuscript that our results were related to the pediatric formulations in use in our hospital. These formulations are “unique” in terms of constituents which, as both Prof. Klek and we stated, included pH, temperature, singular elements and their reactions. At the end of the manuscript, we also stated that “These results may not be extended to other PN admixtures. Therefore further research is warranted to safely act in unstable patients and neonates, who require higher calcium needs and constant variation in nutrients.” The formulations we analyzed came from prescribers working in different Units and we cannot blame Neonatologists if they preferred to keep the calcium content low. From a clinical perspective, neonates did not appear to have side effects despite the low dosage of calcium. Perhaps this was due to their short time on exclusive parenteral nutrition. Admittedly, few prematures had aspects of bone demineralization, but this was partly due to lower than expected phosphorus content released by the source we used which is fructose 1,6diphosphate. Based on our analysis, the critical calcium cut off in parenteral nutrition combined with different lipid emulsions is 4.5 mmol/L. If this limit is confirmed, the gap between it and the recommended dose will further widen. The safety limit of 4.5 mmol/L is particularly critical in combination with fish oil products. However, this does not mean that calcium and fish oil cannot be combined because, in the presence of higher calcium needs, fish oil can still be infused separately. Admixtures compounded with soybean oil, olive oil and soybean-MCToilbased emulsions are stable even if calcium concentration exceeds this limit, as we showed in other analyses [1]. As for the analytical method, we chose the Light Scattering Technique performed by means of a Laser Granulometer to detect lipid particles without aiming to compare it to other techniques. The

Pediatric parenteral admixture compounding in a highly specialized university hospital

Introduction: The aim of this paper is to describe personalized pediatric parenteral admixture compounding in a specialized University Hospital. Materials and methods: Pediatric formulations require small volumes, high concentrations of nutrients and electrolyte micro-additions, thus they must be carefully evaluated by pharmacists and compounded by means of an automatic filling system in a “clean room”. Different kinds of lipid emulsions are used according to the patients’ needs: composite lipids or long and medium chain triglycerides with or without fish oil-based omega-3 fatty acid addition. Admixture stability is defined by determining particle diameter, which must be in the range of 0.4-1.0 µm, like chylomicra, for a safe infusion to patients. Analyses were carried out by means of a laser diffraction technique. Results: In the last 3 years 21,500 personalized pediatric admixtures were compounded (15,480 for children and 6,020 for neonates); the total number of treated patients was 1,700, including 550 neonates. Stability studies pointed out that, in all admixtures, the physicochemical stability did not change between t=0 and t=96 hours and particle diameter was in the expected range of 0.4-1.0 µm provided calcium concentration remained below 9 mEq/L. When calcium exceeded this level, as it is often required for neonates, in the admixtures containing fish oil-based emulsion, 12% of the particles had a diameter larger than 1.0 µm and 2% exceeded 5.0 µm immediately after compounding. Conclusions: Pediatric admixture compounding in a centralized service, and stability studies allowed to prepare any required formulation, providing patients with a safe and effective product. In particular, our analyses pointed out that admixtures containing fish oil-based emulsion and calcium concentrations higher than 9 mEq/L cannot be safely infused to pediatric patients; in these cases it is advisable to infuse the fish oil-based emulsion alone through a second intravenous line.