title: Quantitation of the immunological adjuvants, monophosphoryl lipid A and Quil A in poly (lactic-co-glycolic acid) nanoparticles using high performance liquid chromatography with evaporative light scattering detection.
abstract: Monophosphoryl lipid A (MPL) and Quil A are two immunological adjuvants commonly used in vaccines. At present no simple, validated methods for the quantification of Quil A and MPL have been previously reported therefore the aim of the current study was to develop a simple, fast and validated method to quantify MPL and Quil A using high performance liquid chromatography evaporative light scattering detection (HPLC-ELSD). The HPLC-ELSD technique was carried out using a ZORBAX Eclipse XDB-C8 column (2.1×50 mm; particle size, 3.5 μm) in an isocratic elution mode at 25 °C. MPL was eluted at a retention time of 1.8 min with methanol-water as the mobile phase and a detector temperature of 75 °C. Quil A was resolved as three peaks with retention times of 4.1, 5.5 and 6.4 min with a detector temperature of 30 °C and with water-acetonitrile and 0.01% formic acid as the mobile phase. The nebulizer pressure and gain were set at 3.5 bar and 10, respectively. Calibration curves plotted for both the adjuvants had an R(2)>0.997. Accuracy, intra- and inter-day precision were within the accepted limits. The limit of detection for MPL and Quil A were calculated as 1.343 and 2.06 μg/mL, respectively. The limit of quantification was 2.445 for MPL and 8.97 μg/mL for Quil A. This analytical method was used to quantify the entrapment and in vitro release of MPL and Quil A in a poly lactic-co-glycolic acid (PLGA) nanoparticle vaccine.
authors.affiliation: School of Pharmacy, University of Otago, Dunedin 9054, New Zealand.
authors.forename: Sharan
authors.initials: S
authors.lastname: Bobbala
authors.affiliation: School of Pharmacy, University of Otago, Dunedin 9054, New Zealand.
authors.forename: Arlene
authors.initials: A
authors.lastname: McDowell
authors.affiliation: School of Pharmacy, University of Otago, Dunedin 9054, New Zealand. Electronic address: sarah.hook@otago.ac.nz.
authors.forename: Sarah
authors.initials: S
authors.lastname: Hook
mesh_terms: D000276:Adjuvants, Immunologic / Q000008:administration & dosage / Q000032:analysis*; D002851:Chromatography, High Pressure Liquid / Q000379:methods*; D065546:Drug Liberation; D019344:Lactic Acid / Q000737:chemistry*; D057230:Limit of Detection; D008050:Lipid A / Q000008:administration & dosage / Q000031:analogs & derivatives* / Q000032:analysis; D053758:Nanoparticles / Q000737:chemistry*; D011100:Polyglycolic Acid / Q000737:chemistry*; D000077182:Polylactic Acid-Polyglycolic Acid Copolymer; D062605:Quillaja Saponins / Q000008:administration & dosage / Q000032:analysis*
chemical_list: D000276:Adjuvants, Immunologic; D008050:Lipid A; D062605:Quillaja Saponins; D000077182:Polylactic Acid-Polyglycolic Acid Copolymer; D011100:Polyglycolic Acid; D019344:Lactic Acid; C046386:Quil A; C048436:monophosphoryl lipid A
keywords: Adjuvants; ELSD; Encapsulation efficiency; In vitro release; Nanoparticles; Validation