Development of choline-enhanced formulation for cholorhexidine based antiseptics

Fatemeh Saadatpour,1,*

1. 1 Department of Microbiology, School of Biology and Center of Excellence in Phylogeny of Living Organisms, College of Sc



Effective antisepsis agents are essential in controlling contact transmission of infectious agents within the health-care setting. hospital and laboratory personnel as well as surgeons are forced to use antiseptic compounds daily to minimize the hospital-acquired infection. chlorhexidine gluconate (chg) has been initially introduced by its broad-spectrum antibacterial activity and widely used as a preoperative surgical skin-preparation solution and intra-wound irrigation agent (williamson, carter et al. 2017, liu, werner et al. 2018). however, there are some side effects to chg treatment such as skin inflammation and cytotoxicity effect at high concentrations (liu, werner et al. 2018). with the increase resistance in the prevalence to conventional antiseptics and antibiotics, attention is now turning to the use of natural antibacterial enhancers. choline, as an inherent key in metabolism process, was selected as a candidate for improving the activity of antiseptics. the aim of the work was inyroducing natural and nom-toxic additives that could diminish the common concentration of chg while maintaining or enhancing its anti-microbial effect.


For this purpose, the antimicrobial effects of choline alone and in combination with chg was investigated against standard strain of staphylococcus aureus atcc 6538. killing experiments were performed according to the european standard suspension test une-en-1276 (anonymous, 1998). chg solutions at concentrations of 0.001, 0.005, 0.01, 0.05, 0.1 and 0.2 (v/v) were made in double distil water. the choline solutions was prepared in five concentrations of 0.1,0.3, 0.5, 0.6, 0.7, 1, 1.6, 3.4 and 25 mm. cell suspension was prepared to 107 cfu/ml and were used to validate the assay and to test the disinfectant agent. the bactericidal effect of additive and chlorhexidine against test strain was evaluated using mic, mbc determination based on clsi guideline (patel 2017). the load reduction was evaluated according to the reference standard (une-en-1276) at 30 s, 1, 2, 3, 4, and 5 min. finally, the load and expressed as cfu/ml.


Based on result, choline lonely was shown bactericidal potency against s. aureus atcc 6538 but its effect was less than the concentration of chlorhexidine at 0.001 %. the combination effect of the choline (0.6 mm)–chlorhexidine was more effective against s. aureus atcc 6538. the mic of chlorhexidine was reduced 2-fold in combination with the choline as an additive. the combination of the choline (0.6 mm) –chlorhexidine caused >4 log cfu/ml bacterial load reduction after 2 min of exposure compared to the chlorhexidine and additives lonely at the same times.


Combining chlorhexidine with choline, at a lower level of inhibition, resulted in a substantial reduction in the amount of chlorhexidine required (10-fold reduction) to achieve the same level of growth inhibitionthe antioxidant and anti-inflammatory activity of choline has already been reported but there have been no distinct study describing the antimicrobial effects of it (surai 2015, masisi, beta et al. 2016). the rate of nonsusceptibility to chlorhexidine was also shown an increase in the s. aureus isolates (williamson, carter et al. 2017). present research suggested that augmentation by cholie can lead to declining the cytotoxic effects of excess use of chg in addition to the reduction in chlorhexidine consumption and additionally enhanced formulation might be effective to inhibit the staphylococci strains with resistant to chlorhexidine.


Chlorhexidine gluconate, choline, antiseptic activity