Formulation and stability Optimization of a Choline-Enriched DHA/ARA Emulsion for Preterm Infants

Abstract

Preterm infants benefit from combined supplementation of arachidonic acid (ARA), docosahex-aenoic acid (DHA) and choline, but ARA/DHA-rich oils are highly susceptible to lipid oxida-tion. This project aimed to develop and evaluate choline-enriched DHA/ARA oil-in-water emulsions with suitable physical and oxidative stability. Emulsions were prepared by high-pressure homogenization using two AquaCelle® emulsifier systems which are EOC and GP1, choline chloride and α glycerophosphocholine, and two emulsifier levels. Droplet size evolution over time was used to evaluate physical stability, while peroxide value, conjugated dienes, para-anisidine value and TOTOX were used to assess oxidation during accelerated storage. AUC-based responses were mainly used for formulation comparison. The results showed that emulsi-fier type was the main factor affecting droplet size. EOC produced smaller droplets than GP1 but also showed higher accumulation of secondary oxidation products. In contrast, GP1-containing formulations showed better control of p-AV and overall oxidation burden. Choline form mainly influenced secondary oxidation, with α glycerophosphocholine performing better than choline chloride. Based on Design-Expert screening, GP1 with GPC combination was selected as the most promising formulation, with the highest desirability value.

Popular Abstract

Some nutrients are especially important for very premature infants, but they are also difficult to formulate safely. This project studied how to make an oil-in-water emulsion containing omega-3 and omega-6 and choline, while reducing the risk of oil oxidation. Very premature infants miss part of the nutrient transfer that normally happens during late pregnancy. Among these nutrients, omega-3 and omega-6 are important fatty acids for the developing brain and retina, while cho-line is needed for cell membranes and may help transport these fatty acids in the body. A practi-cal supplement should therefore be able to deliver oil-soluble omega-3 and 6 together with wa-ter-soluble choline in a small and stable dose. One possible solution is an oil-in-water emulsion. This is similar in principle to many everyday products, such as milk or salad dressing: tiny oil droplets are dispersed in water with the help of emulsifiers. Smaller droplets can be good be-cause they may improve stability and digestion. However, there is a challenge. Omega-3 and omega-6 are sensitive to oxidation. When oils oxidize, they can lose nutritional quality and form unwanted breakdown products. For preterm infants, who are already vulnerable to oxidative stress, controlling this process is especially important. In this project, different emulsion formula-tions were prepared by high-pressure homogenization. The formulations differed in emulsifier type (called GP1 and EOC), emulsifier amount and choline form (called CC and GPC). Their droplet size was measured to evaluate stability, and several chemical tests were used to follow oxidation over 15 days. The most interesting finding was that the smallest droplets were not always the best choice. Formulations made with the EOC emulsifier produced smaller droplets, but they also showed more secondary oxidation products. These products appear later in the oxidation process and are more complex than the first oxidation products. In contrast, formula-tions made with the GP1 emulsifier had slightly larger droplets but better control of secondary oxidation. The choline form also mattered. GPC performed better than choline chloride in re-ducing secondary oxidation. Based on the overall screening, the most promising formulation was GP1 with GPC combination. This formulation did not have the smallest droplet size, but it gave the best overall balance for stability. The project shows that formulation development is not only about making droplets as small as possible. For sensitive nutritional emulsions, the sta-bility of the oil is just as important. The selected formulation is not yet a final product, but it provides a useful starting point for further validation, longer storage studies and future bioavail-ability testing

• author: Xue, Ruilin
• supervisor: Marie Wahlgren; Tobias Halthur
• organization: Pharmaceutical Technology (master); Food Technology and Nutrition (M.Sc.)
• course: KLGM06 20261
• year: 2026
• type: H2 - Master's Degree (Two Years)
• subject: Technology and Engineering
• keywords: Oil-in-water emulsion, Emulsifier systems, Physical stability Oxidative stability, Docosahexaenoic acid (DHA), Arachidonic acid (ARA), Choline, Pharmaceutical formulation
• language: English
• id: 9238006
• date added to LUP: 2026-06-16 14:36:47
• date last changed: 2026-06-16 14:36:47