High-performance size-exclusion chromatographic procedure for the determination of fluoresceinyl isothiocyanate dextrans of various molecular masses in biological media

https://doi.org/10.1016/S0378-4347(00)82825-XGet rights and content

Abstract

A high-performance size-exclusion chromatographic procedure, using Nucleosil Diol, for the quantitative analysis of fluoresceinyl isothiocyanate dextrans of various molecular masses (10 000–150 000) in biological media was developed. The influence of the molecular mass and the degree of substitution of the conjugates on the chromatographic behaviour are discussed. In addition to quantitation, the molecular mass of the conjugates with degree of substitution below 1.6 could be estimated from the chromatograms. Linear standard calibration curves were obtained at concentrations down to 0.050 μg ml−1 in rabbit plasma and urine and homogenates of rabbit liver, lymph node and muscle when the derivative (degree of substitution 0.85) was monitored by fluorescence detection (λex=493 nm, λem=520 nm). The fluoresceinyl isothiocyanate dextrans were found to be stable for more than three days at 37°C in all biological media under investigation. A pH-rate profile for the alkaline hydrolysis of fluoresceinyl isothiocyanate dextrans was constructed. The applicability of the method to pharmacokinetic studies was demonstrated by recording the plasma concentration-time profile of a fluoresceinyl isothiocyanate dextran T-70 conjugate following intravenous injection to a rabbit. In relation to future pharmacokinetic investigations on dextran conjugates, the results reported indicate that labelling of the parent dextran with fluoresceinyl isothiocyanate and monitoring of the fluoresceinyl isothiocyanate dextran conjugate throughout the organism using the described method is a promising development.

References (26)

  • R.G. Melton et al.

    Biochem. Pharmacol.

    (1987)
  • N.R. Worrell et al.

    Biochem. Pharmacol.

    (1986)
  • J.C. Glover et al.

    J. Neurosci. Methods

    (1986)
  • J.P. Camilleri et al.

    Exp. Mol. Pathol.

    (1983)
  • J.A. Oliver et al.

    Methods Enzymol.

    (1984)
  • U. Schröder et al.

    Microvasc. Res.

    (1976)
  • A.N. de Belder et al.

    Carbohydr. Res.

    (1973)
  • C. Larsen et al.

    J. Chromatogr.

    (1987)
  • G. Sartore et al.

    Tetrahedron Lett.

    (1974)
  • L.W. Dittert et al.

    J. Pharm. Sci.

    (1963)
  • C. Larsen et al.

    Arch. Pharm. Chem.

    (1985)
  • H. Sezaki et al.

    CRC Crit. Rev. Ther. Drug Carrier Systems

    (1984)
  • M. Hashida et al.

    Cancer Res.

    (1984)
  • Cited by (21)

    • Assay considerations for fluorescein isothiocyanate-dextran (FITC-d): an indicator of intestinal permeability in broiler chickens

      2021, Poultry Science
      Citation Excerpt :

      For 4-kDa FITC-d, the fluorescence measurement is conducted with a microplate reader at an excitation wavelength of 485 nm and the emission wavelength of 528 nm (Kuttappan et al., 2015ab). A previous study reported that FITC-d were stable for more than three days at 37°C in biological media (Kurtzhals et al., 1989). In poultry studies, some researchers conducted the FITC-d measurements on the same day of the sampling (Bortoluzzi et al., 2019a; Teng et al., 2020), whereas other researchers have kept serum samples at –20 C until further analysis (Gilani et al., 2017b).

    • Vitreal elimination kinetics of large molecular weight FITC-labeled dextrans in albino rabbits using a novel microsampling technique

      2000, Journal of Pharmaceutical Sciences
      Citation Excerpt :

      A 250‐mm × 4.6‐mm Microsphere GPC 150 size exclusion column (7 μ) obtained from Altech Associates (Deerfield, IL) was used for the quantitation of the vitreous and aqueous samples. The size exclusion analytical method was adapted from Kuetzhals et al15 with modifications. The mobile phase consisted of 0.01 M ammonium phosphate buffer (pH 7.0).

    View all citing articles on Scopus
    View full text