March 2012
Volume 53, Issue 14
Free
ARVO Annual Meeting Abstract  |   March 2012
Demonstration of Naturally Occurring Lacritin as an Antimicrobial Preservative in Topical Ophthalmic Preparations
Author Affiliations & Notes
  • Evan M. Berger
    TR Lee Center for Ocular Pharmacology, Eastern Virginia Medical School, Norfolk, Virginia
  • Sandeep S. Samudre
    TR Lee Center for Ocular Pharmacology, Eastern Virginia Medical School, Norfolk, Virginia
  • Alireza Hosseini
    TR Lee Center for Ocular Pharmacology, Eastern Virginia Medical School, Norfolk, Virginia
  • Patricia B. Williams
    TR Lee Center for Ocular Pharmacology, Eastern Virginia Medical School, Norfolk, Virginia
  • Frank A. Lattanzio, Jr.
    TR Lee Center for Ocular Pharmacology, Eastern Virginia Medical School, Norfolk, Virginia
  • Footnotes
    Commercial Relationships  Evan M. Berger, None; Sandeep S. Samudre, EyeRx Inc. (R); Alireza Hosseini, None; Patricia B. Williams, EyeRx Inc. (R); Frank A. Lattanzio, Jr., EyeRx Inc. (R)
  • Footnotes
    Support  NEI #1R41EY020044-01
Investigative Ophthalmology & Visual Science March 2012, Vol.53, 1860. doi:
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      Evan M. Berger, Sandeep S. Samudre, Alireza Hosseini, Patricia B. Williams, Frank A. Lattanzio, Jr.; Demonstration of Naturally Occurring Lacritin as an Antimicrobial Preservative in Topical Ophthalmic Preparations. Invest. Ophthalmol. Vis. Sci. 2012;53(14):1860.

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      © ARVO (1962-2015); The Authors (2016-present)

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Abstract

Purpose: : Lacritin is a novel glycoprotein found in tears that has shown potential as an alternative paradigm in antimicrobial therapy. Benzalkonium chloride (BAK), currently the most widely used preservative in topical ophthalmic preparations, is cytotoxic and damages the human cornea. We propose that lacritin, as a naturally occurring protein in human tears, could act as an antimicrobial preservative when formulated into solution.

Methods: : Test solutions of either N65 lacritin (224, 112, or 56 µg/mL) or BAK (200, 100, or 50 µg/mL), were prepared in Phosphate Buffered Saline (PBS). A negative control contained PBS alone. Cultures of P. aeruginosa (ATCC# 9027) were grown in stock cultures using Luria-Bertani broth or agar. An inoculum was diluted to obtain 105-106 bacteria/mL. Each test solution was then challenged with P. aeruginosa and incubated at 37ºC for 28 days. To determine efficacy, a 100 µL aliquot from each test tube was plated at baseline, 6 and 24 hours, and 7, 14, and 28 days, and then incubated for 24 hours at 37ºC. Following each incubation, the total colony forming units (CFU) were counted on each plate. A preservative efficacy test requires at least a 1.0 log reduction in CFU by the 7th day. Data is presented as mean ± SD.

Results: : Although initially within 6 hours after N65 (112 ug/ml) treatment bacterial colonies were reduced by one log unit, from a baseline of 133±10 to 21±5 CFU, this effect was not sustained. There was no log reduction in CFU after PBS treatment within 6 hours. A baseline of 166±25 CFU after 6 hours was similar to 296±18 CFU at baseline (p>0.05). The CFU after treatment with either N65 or PBS were too numerous to count after 24 hours as the plates became confluent. The study was terminated after 7 days because there was no further reduction in CFU. In contrast to N65 and PBS treatment, the BAK treatment was most efficacious as there was no observed growth in CFU at any time point in the study.

Conclusions: : Using the criteria for the preservative efficacy test [USP 29(2006)], we found BAK to be more efficacious than N65. Being a biological compound, N65 may be unstable in vitro and therefore lose activity by 24 hours. Although initially effective as a preservative, N65 may be beneficial as an adjuvant because it has the ability to preserve and restore the corneal surface and therefore negate the untoward effects of BAK. Future investigations will focus on using N65 as a corneal surface protectant.

Keywords: bacterial disease • ocular irritants • drug toxicity/drug effects 
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