Ocular Pharmacokinetics of 25-Hydroxyvitamin D3 After Weekly Supplementation in Rabbits Using Ultra Performance Liquid Chromatography–Tandem Mass Spectrometer

  • Archit Kumar
  • Mini P. Singh
  • Tripti Rungta
  • Ajay Patial
  • Savita Verma Attri
  • Bikash Medhi
  • Radha Kanta Ratho
Short Communication


Background and Objectives

The protective role of vitamin D supplementation has recently been shown to be present in various ocular inflammatory diseases. The oral supplementation of vitamin D may take time to achieve adequate levels in intraocular fluids. Therefore, the present study was performed to understand the ocular pharmacokinetics of 25-hydroxyvitamin D3 (25D3) in aqueous humor after weekly supplementation of 25D3 in rabbits.


A total of 21 rabbits were fed orally with 25D3 (7.22 µg/kg/week) for 8 weeks and 9th dose was given at the end of 8 weeks. The blood and aqueous humor samples were collected from ear vein and though anterior chamber paracentesis, respectively. The serum and aqueous humor samples were spiked with deuterium labeled internal standard and were extracted using liquid extraction method. Furthermore, the samples were derivatized and 25D3 estimation was performed using ultra performance liquid chromatography–tandem mass spectrometer (UHPLC–MS/MS).


The 25D3 supplementation significantly increased the 25D3 levels in serum (78.5 ± 21.6 ng/ml) (mean ± SD) (p < 0.0001) and in aqueous humor (991.3 ± 180.6 pg/ml) (mean ± SD) (p < 0.0001) compared to baseline levels. The maximum concentration was achieved in serum after the 10th hour of supplementation of 1st and 9th dose, while the same was observed at the 24th hour in aqueous humor.


The oral supplementation of 25D3 was found to significantly increase 25D3 levels in aqueous humor; however, the time required to achieve 25D3 concentration in aqueous humor was higher as compared to that in serum. Therefore, weekly oral supplementation of 25D3 may have a beneficial role in ocular diseases.



We thank Shefali Saini for her assistance in experiments when required.

Author Contributions

AK, MPS and BM conceived the idea and planned the study design. The experiments were performed by AK. The LCMS were performed by AP and AK. TR assisted during animal experiments and LCMS. The paper was drafted by AK and MPS and critically reviewed by SV, BM and RKR.

Compliance with ethical standards

Conflict of interest

The authors have declared no conflict of interest.

Ethical approval

The animal experimentations were performed after obtaining animal ethical clearance from Institute Animal Ethics Committee (Reference No. IAEC-66/IAEC/396).


The study was funded by Department of Biotechnology, New Delhi Reference No. BT/PR10811/MED/29/840/2014.


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Copyright information

© Springer International Publishing AG, part of Springer Nature 2018

Authors and Affiliations

  1. 1.Department of VirologyPostgraduate Institute of Medical Education and ResearchChandigarhIndia
  2. 2.Advanced Pediatrics CentrePostgraduate Institute of Medical Education and ResearchChandigarhIndia
  3. 3.Department of PharmacologyPostgraduate Institute of Medical Education and ResearchChandigarhIndia

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