Journal of Food Hygiene and Safety (한국식품위생안전성학회지)

The Korean Society of Food Hygiene and Safety (한국식품위생안전성학회)
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Comparison of In Vitro, Ex Vivo, and In Vivo Antibacterial Activity Test Methods for Hand Hygiene Products

손 위생 제품에 대한 in vitro, ex vivo, in vivo 항균 시험법 비교

  • 이다은 (코스맥스 R&I센터, 안전성 랩) ;
  • 여현주 (코스맥스 R&I센터, 안전성 랩) ;
  • 정혜윤 (코스맥스 R&I센터, 안전성 랩)
  • Received : 2023.10.26
  • Accepted : 2024.01.08
  • Published : 2024.02.28
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Abstract

Numerous methods have been applied to assess the antibacterial effectiveness of hand hygiene products. However, the different results obtained through various evaluation methods have complicated our understanding of the real efficacy of the products. Few studies have compared test methods for assessing the efficacy of hand hygiene products. In particular, reports on ex vivo pig skin testing are limited. This study aimed to compare and characterize the methodologies applied for evaluating hand hygiene products, involving in vitro, ex vivo, and in vivo approaches, applicable to both leave-on sanitizers and wash-off products. Our further aim was to enhance the reliability of ex vivo test protocols by identifying influential factors. We performed an in vitro method (EN1276) and an in vivo test (EN1499 and ASTM2755) with at least 20 participants, against Serratia marcescens or Escherichia coli and Staphylococcus aureus. For the ex vivo experiment, we used pig skin squares prepared in the same way as those used in the in vivo test method and determined the optimal treated sample volumes for sanitizers and the amount of water required to wash off the product. The hand sanitizers showed at least a 5-log reduction in bacterial load in the in vitro test, while they showed little antibacterial activity in the in vivo and ex vivo tests, particularly those with a low alcohol content. For the hand wash products, the in vitro test was limited because of bubble formation or the high viscosity of the products and it showed low antibacterial activity of less than a 1-log reduction against E. coli. In contrast, significantly higher log reductions were observed in ex vivo and in vivo tests, consistently demonstrating these results across the two methods. Our findings revealed that the ex vivo and in vivo tests reflect the two different antibacterial mechanisms of leave-on and wash-off products. Our proposed optimized ex vivo test was more rapid and more precise than the in vitro test to evaluate antibacterial results.

손 위생 제품이 다양화됨과 동시에 각 활용 방법에 따라 그 효능을 평가하는 여러 시험 방법들이 보고되고 있다. 하지만 평가 방법에 따라 각 제품의 항균 효능은 다르게 나타나며, 이로 인해 제품의 실제적인 효능을 확인하는 데에 어려움이 있을 수 있다. 손 위생 제품의 효능평가방법 비교에 초점을 둔 연구는 매우 제한적이며, 특히 돼지피부를 이용한 ex vivo에 대한 연구는 극히 드물다. 이에 본 연구는 손 위생 제품 중 리브온 소독제와 워시오프 세정제에 대해 각각의 항균 평가 방법을 종합적으로 비교했고, ex vivo 시험에 영향을 미칠 수 있는 요인을 파악하여 연구 단계에서 효율적인 ex vivo 시험의 신뢰성을 향상시키고자 하였다. in vitro 시험으로써 액체 현탁을 기반으로 하는 time-kill 시험을 진행했고, in vivo 시험은 최소 20명의 참여자를 대상으로 진행되었다. ex vivo 시험은 규격화된 돼지 피부를 이용하여 in vivo 시험과 동일한 방법으로 진행하면서 소독제의 최적 처리량과 세정제 사용 시 첨가되는 물의 양을 제안했다. 시험에 사용된 손 소독제는 in vitro 시험에서 모두 5 log 이상의 세균 감소율을 보인 반면, ex vivo와 in vivo에서는 훨씬 낮은 살균 활성을 보였으며, 특히 알코올 함량이 낮은 손 소독제에서는 1 log 미만의 살균 활성을 나타냈다. 반면에 손 세정제의 in vitro 시험 결과, 대장균에 대해서는 1 log 이하의 낮은 항균력을 보였으나, ex vivo 와 in vivo 시험 결과에서는 이보다 높은 항균력을 유사하게 나타냈다. 본 연구에서는 ex vivo 와 in vivo 시험 방법이 리브온과 워시오프 타입 제품의 두가지 다른 항균 메커니즘을 반영할 수 있음을 확인했다. 이로 인해 최적의 조건으로 설정된 ex vivo 시험은 빠르고 정확한 항균 평가법이 될 수 있음을 제시한다.

Keywords

Acknowledgement

We would like to express our gratitude to the Korea Testing & Research Institute and the Global Medical Research Center for assistance with data analysis.

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