Water soluble inner aqueous phase markers as indicators of the encapsulation properties of water-in-oil-in-water emulsions stabilized with sodium caseinate

The aim of this study was to evaluate the suitability of Methylene Blue (MB) and Vitamin B₁₂ (Vit-B₁₂) as water soluble inner aqueous phase (W₁) markers for measuring the encapsulation efficiency and stability of water-in-oil-in-water (W₁/O/W₂) double emulsions stabilized by sodium caseinate (NaCN). The encapsulation efficiency and stability were determined by centrifugation of the double emulsion to separate the cream phase (W₁/O) and the outer aqueous phase (W₂) and measuring the concentration of marker in W₂ by absorbance spectrophotometry. To validate this method the marker concentration measurable and the stability of the marker in W₂ were measured. Both markers could be accurately measured in W₂ and there was no change in the concentration of marker on storage of a W₂ solution for 7 days at 45 °C. The recovery yields of MB and Vit-B₁₂ in the recovered W₂ of an oil-in-water (O/W₂) emulsion, determined using the procedure normally used for measuring encapsulation efficiency and stability, were 78% and 99%, respectively, and 52 and 100%, respectively. Double emulsions had encapsulation efficiency of 61.9 ± 21.4% and 16.6 ± 1.1% and encapsulation stability of 62.0 ± 22.6% and 10.7 ± 0.7% for MB and Vit-B₁₂, respectively. Recovery yield and encapsulation efficiency/stability data for MB indicate that it is not a suitable marker for measuring the encapsulation properties of NaCN stabilized double emulsions while similar data for Vit-B₁₂ indicate that it is a suitable marker for studying the encapsulation properties of double emulsions stabilized with NaCN. Methods used in other studies to measure encapsulation properties of double emulsions are discussed in light of the results obtained in this study.