Anti-Infective Agents in Medicinal Chemistry, 2008, 7, 281-284 281 1871-5214/08 $55.00+.00 © 2008 Bentham Science Publishers Ltd. Microbiological Efficacy of Throat Antiseptics Used as A Spray or A Solution in Healthy Volunteers N.O. Huebner 1 , O. Assadian 2, *, I. Harfenstein 3 , R. Moesges 4 , T. Koburger 3 , U. Weber 5 and A. Kramer 1 1 Institute for Hygiene and Environmental Medicine, Ernst Moritz Arndt University, Walther-Rathenau-Str. 49 a, 17489 Greifswald, Germany; 2 Clinical Institute for Hygiene and Medical Microbiology, Vienna General Hospital, Medical University of Vienna, Waehringer Guertel 18-20, A - 1090 Vienna, Austria; 3 Hygiene Nord GmbH, Walther-Rathenau- Str. 49 a, 17489 Greifswald, Germany; 4 Institute of Medical Statistics, Informatic and Epidemiology, University of Co- logne, Lindenburger Allee 42, 50931 Cologne, Germany; 5 ANTISEPTICA Medical Scientific, Fischstraße 24a, 17489 Greifswald, Germany Abstract: The aim of this study was to investigate the efficacy different of throat antiseptics on the oropharynx in healthy volunteers. A newly developed standardised test method was established. Using 12 volunteers per product to be tested, the reduction in the total colony count was the primary endpoint, and reduction in viridans Streptococci was the secondary endpoint criterion, for the commercial antiseptics neo-angin® N throat lozenges, neo-angin® N sugar-free lozenges, neo- angin® N throat and mouth spray, Sorot® lozenges, and Hexoral ® mouth wash. At baseline, immediately, 2 and 5 hours after application two areas of the oral mucosa in the lower jaw at the level of the second molars and from the uvular re- gion were swabbed and investigated for total bacterial counts. Following neutralisation, the swabs were inoculated onto blood agar. The antiseptic effect was more prominent in the throat, the area targeted by the antiseptic, than in the cheek mucosa. The reduction in total colony count was significant both with the test preparations and with the comparison preparations; A reduction of > 1 log was achieved for both endpoint criteria immediately after application of antiseptic and continued at this level for 30 minutes. Even after 5 hours, a significant reduction in total colony count was observable. Furthermore the established test model allowed determining the antiseptic efficacy of lozenges or spraying preparations in comparison to rinsing solutions. Keywords: Antiseptics, throat, antimicrobial, methodology, Viridans streptococci, dichlorobenzyl alcohol, amylmetacresol, menthol, dequalinium chloride, hexetidine. INTRODUCTION The aim of this study was to investigate whether and to which extent the pharmaceutical formulations of three throat antiseptics, neo-angin ® N, neo-angin ® N sugar-free and neo- angin ® N throat and mouth spray, cause a reduction in mi- croorganisms within the oropharynx compared to two refer- ence products, Sorot® lozenges and Hexoral ® mouth wash, in healthy volunteers. The following indication has been declared by the manufacturer for these antiseptics: Suppor- tive treatment of inflammations of the mucous membranes of the pharyngeal region accompanied by typical symptoms such as sore throat, redness or swelling. The total colony count was selected as the primary end- point criterion, since this encompasses the microorganisms of the oral and pharyngeal cavities that can be cultured in aerobic conditions on blood agar, and thus can be considered to be representative. Viridans Streptococci were defined as the secondary endpoint criterion, because these are signifi- cant from the viewpoint of infection [1] and are possibly suitable as indicator organisms. *Address correspondence to this author at the Clinical Institute for Hygiene and Medical Microbiology, Medical University Vienna, General Hospital Vienna, Waehringer Guertel 18-20, 1090 Vienna, Austria; Tel: +43-1-404 00-1901; Fax: +43-1-40400-1907; E-mail: ojan.assadian@meduniwien.ac.at A pilot study conducted prior to the investigation pro- vided information about the practicability of the study strat- egy, indications of the expected range of values and data for calculation of sample size. MATERIALS AND METHODS Pilot Study Using the same study design as below, the variation in the reduction factor was determined in 3 subjects with neo- angin® N throat lozenges and neo-angin® N throat and mouth spray, in order to calculate the sample size for the actual study. Test Substances The test products had the following composition: neo- angin® N throat lozenges and neo-angin® N sugar-free loz- enges: 1.2 mg 2,4-dichlorobenzyl alcohol, 0.6 mg amyl- metacresol and 5.9 mg (or 5.72 mg, respectively) men- thol/lozenge; neo-angin ® N throat and mouth spray: 14.58 mg 2,4-dichlorobenzyl alcohol, 2.92 mg amylmetacresol and 0.87 mg menthol/100 ml (Divapharma GmbH, Germany). As a comparison product for the lozenges, a lozenge containing 0.25 mg dequalinium chloride was produced by the phar-