[Toxicity of a new moistening agent and preservative in vitro].
OBJECTIVE
The use of preservatives such as benzalkonium chloride (BAC) usually increases the toxicity of pharmaceutical tear substitutes. HP-guar has been recently introduced as a new artificial tear substitute and includes the preservative Polyquad (0.001%), which is considered to be non-toxic. We therefore examined the effect of preserved (cetrimide 0.01%) and unpreserved HPMC (hydroxypropylmethyl cellulose) and HP-guar in dose and time-response experiments in a human corneal and conjunctival epithelial cell culture model.


METHODS
Immortalized human conjunctival and corneal epithelial cells were cultured in 96-well plates at 37 degrees C with 5% CO(2) and exposed to the test solutions. The ATP content was quantified by means of a luminescence-based ATP assay, intracellular esterase activity by double fluorescent viability staining (calcein AM/ethidium homodimer D-1) and cell migration by a colony dispersion assay. All experiments were performed in triplicate and repeated at least once. The significance of differences was determined with an unpaired two-sided t-test.


RESULTS
HPMC with preservative severely reduced the ATP content at all concentrations tested. Unpreserved HPMC, however, showed an inhibition of ATP production only at 100% and good esterase activity. HP-guar with and without preservative were found to reduce ATP activity more than unpreserved HPMC, but the unpreserved solution was found to reduce cellular ATP levels significantly more than the preserved solution.


CONCLUSIONS
The new preservative Polyquad induced significantly less cytotoxicity than cetrimide. However, even unpreserved HP-guar can induce cytotoxicity in vitro, while unpreserved HPMC remains a good alternative tear substitute with low cytotoxicity.
Kasper
K
K
Augen- und Poliklinik, Universität Würzburg, Josef-Schneider-Strasse 11, 97080, Würzburg, Deutschland.
Kremling
C
C
Geerling
G
G
D000255:Adenosine Triphosphate / Q000378:metabolism; D002465:Cell Movement / Q000187:drug effects*; D002470:Cell Survival / Q000187:drug effects*; D002478:Cells, Cultured; D000077286:Cetrimonium; D002593:Cetrimonium Compounds / Q000633:toxicity*; D003228:Conjunctiva / Q000187:drug effects*; D018639:Contact Lens Solutions / Q000633:toxicity*; D004305:Dose-Response Relationship, Drug; D004847:Epithelial Cells / Q000187:drug effects*; D019573:Epithelium, Corneal / Q000187:drug effects*; D004950:Esterases / Q000378:metabolism; D006801:Humans; D006863:Hydrogen-Ion Concentration; D065347:Hypromellose Derivatives; D066298:In Vitro Techniques; D008747:Methylcellulose / Q000031:analogs & derivatives* / Q000633:toxicity; D009883:Ophthalmic Solutions / Q000633:toxicity*; D009994:Osmolar Concentration; D011108:Polymers / Q000633:toxicity*; D011134:Polysaccharides / Q000633:toxicity*; D011310:Preservatives, Pharmaceutical / Q000633:toxicity*
D002593:Cetrimonium Compounds; D018639:Contact Lens Solutions; D009883:Ophthalmic Solutions; D011108:Polymers; D011134:Polysaccharides; D011310:Preservatives, Pharmaceutical; C519976:hydroxypropyl guar; D065347:Hypromellose Derivatives; C058655:polyquaternium 1; D000255:Adenosine Triphosphate; D008747:Methylcellulose; D004950:Esterases; D000077286:Cetrimonium