AAPS PharmSciTech

, Volume 18, Issue 5, pp 1595–1604 | Cite as

Analytical Characterization of an Oil-in-Water Adjuvant Emulsion

  • Jenny Sun
  • Richard L. RemmeleJr
  • Gautam Sanyal
Research Article


Adjuvants are typically used in subunit vaccine formulations to enhance immune responses elicited by individual antigens. Physical chemical characterization of novel adjuvants is an important step in ensuring their effective use in vaccine formulations. This paper reports application of a panel of quantitative assays developed to analyze and characterize an oil-in-water adjuvant emulsion, which contains glucopyranosyl lipid A (GLA) and is a squalene-based emulsion. GLA is a fully synthetic analogue of monophosphoryl lipid A, which is a Toll-like receptor type 4 agonist and an FDA-approved adjuvant. The GLA-stable emulsion (GLA-SE) is currently being used for a respiratory syncytial virus vaccine in a phase 2 clinical trial. GLA was quantitated using reverse-phased high-performance liquid chromatography (RP-HPLC) coupled to a mass spectrometric detector, achieving higher assay sensitivity than the charged aerosol detection routinely used. Quantitation of the excipients of GLA-SE, including squalene, egg phosphatidyl choline, and Poloxamer 188, was achieved using a simple and rapid RP-HPLC method with evaporative light scattering detection, eliminating chemical derivatization typically required for these chromophore-lacking compounds. DL-α-tocopherol, the antioxidant of the GLA-SE, was quantitated using a RP-HPLC method with conventional UV detection. The experimental results compared well with values expected for these compounds based on targeted composition of the adjuvant. The assays were applied to identify degradation of individual components in a GLA-SE sample that degraded into distinct aqueous and oil phases. The methods developed and reported here are effective tools in monitoring physicochemical integrity of the adjuvant, as well as in formulation studies.

Key words

adjuvant evaporative light scattering detection (ELSD) excipient glucopyranosyl lipid A liquid chromatography with mass spectrometry (LC-MS) vaccine 



The authors like to thank the NanoImaging Services (San Diego, CA) for providing the cryo-EM data.

Compliance with Ethical Standards

Conflict of Interest

The authors declare that they have no conflict of interest.

Supplementary material

12249_2016_626_MOESM1_ESM.docx (875 kb)
Esm 1 (DOCX 875 kb)


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Copyright information

© American Association of Pharmaceutical Scientists 2016

Authors and Affiliations

  • Jenny Sun
    • 1
  • Richard L. RemmeleJr
    • 1
  • Gautam Sanyal
    • 1
  1. 1.Analytical Biotechnology, MedImmune Inc.GaithersburgUSA

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