Persistent and Residual Antimicrobial Effects: Are They Important in the Clinical Setting?

Persistent and Residual Antimicrobial Effects: Are They Important in the Clinical Setting?

By Daryl S. Paulson, PhD

Antiseptics marketed as surgical scrub formulationsmust meet certain effectiveness requirements that relate to their immediate,persistent, and preferably, residual antimicrobial properties. Immediateantimicrobial properties are the antimicrobiocidal effect on residentmicroorganisms immediately (within one minute) after the antiseptic has beenapplied. Generally, this is measured as reductions from the baseline microbialpopulation present on the skin of the hands prior to product application. Thepersistent antimicrobial effect measures how long after its application anantiseptic is able to prevent microbial counts from exceeding the baselinepopulation numbers. This is a severe challenge for surgical scrub/wash products,in that the hands are occluded inside surgeons gloves following productapplication. Occlusion presents a favorable environment for microbial growth.Residual antimicrobial effects measure the products cumulative antimicrobialeffects when used repeatedly over a number of days; that is, someantimicrobials, when used over time, are adsorbed to the stratum corneum of theskin and, as a result, will prevent microbial re-colonization of the skinsurfaces to baseline levels.

The Food and Drug Administrations (FDA) specific test requirements must bemet in order to label the product as a surgical scrub. Although in vitro antimicrobialefficacy must be demonstrated in time-kill kinetic and minimum inhibitoryconcentration studies, the focus of this paper is in vivo simulated humanclinical trials. The FDA surgical scrub evaluation is four weeks in duration. Itbegins with a two-week wash out period, during which subjects do not useany antimicrobial products so that the population numbers of their normal handflora are stabilized. It is followed by a one-week baseline period, during whichthe normal number of microorganisms per hand is measured. All the subjectsbaseline measurement data are pooled to provide a baseline point-estimate valueto which the post-product use values are compared. The next week (week four) is the test week, when the antiseptic is usedeleven times over the course of five days, per label instructions. Table 1provides the product application and sampling schema.

To meet FDA label claim requirements, the antiseptic must produce at least aone log10 reduction from baseline population on test day one, at least a 2 log10reduction from baseline population on test day two, and at least a 3 log10reduction from baseline population on test day five. Further, the microbial populations of the gloved hands cannot exceed baselinepopulation over the six-hour post-application period on any of the threeevaluation days.

Figure 1 depicts a surgical scrub formulation that meets the FDArequirements, starting with a baseline population at a log10 value of five, theminimum permitted by the FDA for testing of surgical scrub antiseptics. Thefigure illustrates plainly the immediate, persistent and residual antimicrobialeffects of the product.

Table 1. Test Week Product Application and Sampling Schema

 1 product application followed by sampling immediate, 3-hours and 6-hours post-product application1 product application followed by sampling immediate, 3-hours and 6-hours post-product application3 product applications, at least 1 hour apart3 product applications, at least 1 hour apart1 product application followed by sampling immediate, 3-hours and 6-hours post-product application
 no additional applications2 additional surgical product applications, at least 1 hour apartno samplesno samplesno additional applications
 1 total product application3 total product applications3 total product applications3 total product applications1 total product application

Figure 2 portrays the results from the testing of an antiseptic that did notmeet the FDAs requirements for immediate antimicrobial reductions on any testday nor day ones persistent effectiveness criterion, because the microbialcount at six hours exceeds the baseline counts.

Whether an antiseptic meets the FDA requirements, to a large degree, dependson the active ingredient and its concentration, and on the overall productformulation. Table 2 presents the performance of common antimicrobial activesthat is typically observed, when they are used as surgical scrub handantiseptics.

Table 2. Activities of Common Antimicrobial Chemicals

 Antimicrobial Properties
70% Isopropyl AlcoholHighNone*None
4% Chlorhexidine Gluconate (CHG)HighHighHigh
Povidone IodineModerateModerateNone
Alcohol and Povidine IodineHigh**HighNone
Alcohol and ChlorhexidineGluconate (CHG)HighHighHigh

** Alcohol (60 percent to 90 percent) generally increases thespeed of microbial killing of both chlorhexidine gluconate and povidone iodine.

Recall that persistent and residual antimicrobial effects can be achieved bymerely inhibiting microbial growth of normal flora,;a passiveantimicrobial effect. So, for hands contaminated with microorganisms subsequentto product use, the FDA has no requirement that the treated hands demonstrateany antimicrobial effects. Yet, one of the main arguments for handwashing orscrubbing with an effective antiseptic is to provide not only persistent and/orresidual activity that maintains microbial counts of normal flora below baselinelevels, but also to provide active extended antiseptic effectivenessagainst contaminating microorganisms. But do any of the commonly used topicalantimicrobials provide active properties that actually prevent growth of or killmicroorganisms that contaminate the hands after they have been scrubbed? Toanswer this question, six types of antiseptic products were evaluated in thisstudy.

Methods and Materials

A simple, visual test was designed to demonstrate active and residualproperties of six antiseptic products, if they were present. The antimicrobialactives used in this test were:

  • 4 percent Chlorhexidine gluconate (CHG)

  • 15 percent Povidone Iodine(PVP-I) Cleansing Solution

  • 3.3 percent Parachlorometaxylenol (PCMX) Emollient Cleansing Solution

  • 61 percent Ethyl Alcohol 70 percent Isopropyl Alcohol

  • 1 percent CHG in 61 percent Ethyl Alcohol


Step 1. The seven days prior to the commencement ofthe evaluation constituted the wash-out period.

Step 2. On test day one, subjects, with the assistance of a trainedtechnician, placed their hands, palm-side down, firmly onto agar platesinoculated with Staphylococcus aureus (ATCC #6538). The hands remainedpressed onto the plates for 60 seconds. This constituted the no-treatment,or baseline measurement.

Step 3. Following the baseline hand-imprinting procedure, the subjectswashed their hands with their assigned product and again pressed their handsonto inoculated agar plates, as described in Step 2. This constituted an immediate antimicrobial sampling, following whichsubjects applied their assigned antiseptic product two additional times beforebeing allowed to leave the laboratory.

Step 4. On test days two through five, subjects returned to thelaboratory to apply their assigned product to their hands three consecutivetimes on each day.

Step 5. Approximately one hour and six hours after the final productapplication on test day five, subjects again performed the hand-imprintingprocedure, as described in Step 2.

Step 6. The hand-imprinted agar plates were incubated at 35 degrees ± 2degrees C for 24 to 48 hours.

Step 7. The agar plates imprinted for baseline and post-productapplication on days one and five were photographed, following incubation. They provided the growth/no growth data to be evaluated visually.


Photograph 1 shows a representative inoculated agar plate touched by anuntreated hand.

A hand imprint can be observed, but the Staphylococcus aureus withinthe imprint outline obvious was neither killed nor inhibited by the untreatedhand.

A hand imprint six hours after the application of the 4 percent CHG producton day five, as displayed in photograph 2, plainly provided a high degree ofantimicrobial activity. That is, the antimicrobial was actually transferred to the plate, preventingmicrobial growth over the period of incubation. This effect was the result of both persistent and residual activity of theCHG.

A hand imprint six hours after the use of the 15 percent PVP-I product on dayfive produced essentially no active antimicrobial effects (photograph 3).In fact, bacterial proliferation within the hand print is indistinguishable fromthat for the hand imprinted for baseline (Photograph 1).

Essentially no active antimicrobial effects are noted for a hand treated withthe 3.3 percent PCMX product at the six-hour post-product application samplingtime on day five (Photograph 4).

Neither of the alcohols, 61 percent ethyl and 70 percent isopropyl, producedany antimicrobial effect on the Staphylococcus aureus six hours afterapplication on day five (Photographs 5 and 6). This was as expected, becauseonce alcohols dry, they no longer have any antimicrobial properties.

The 1 percent tincture of CHG, at six hours post-application on day five,showed a high degree of antimicrobial effectiveness, although less than expected(Photograph 7).

This result likely was due less to a lack of persistent and residualproperties in the CHG, itself, but rather to some blocking by emollientscontained in the product formulation.


This evaluation clearly demonstrated visually that some antiseptics willremain antimicrobially active on the hands, able not only to keep microbialpopulations of normal flora below baseline levels, but also to kill or inhibitmicroorganisms contacted by the treated hands. Perhaps, then, it is reasonableto refer to two types of persistent and residual antimicrobial properties, asopposed to only latently passive activity that keeps microorganisms on thehands below baseline counts. The second type of persistent/residualantimicrobial effects, which this study has graphically illustrated, can betermed latently active. This is because these products not only provide latentpassive protection, but they also show persistent activity againstcontaminative bacteria for an extended period after use. Such a property assuredly is of value in the medical arena.

Daryl S. Paulson, PhD, is with Bozeman, Montana-based BioScienceLaboratories, Inc.

Skin Antisepsis Basics

The recommendations from the CDC Guideline for Hand Hygiene in HealthcareSettings place great emphasis on the thorough decontamination of healthcareworkers hands. The guidelines, in part, recommend the performance of surgical handantisepsis using either an antimicrobial soap or an alcohol-based hand rub withpersistent activity before donning sterile gloves when performing surgicalprocedures. When performing surgical hand antisepsis using an antimicrobialsoap, healthcare workers should scrub their hands and forearms for the length oftime recommended by the manufacturer, usually two to six minutes. When using analcohol-based surgical hand scrub product with persistent activity, healthcareworkers should follow the manufacturers instructions. Before applying thealcohol solution, pre-wash hands and forearms with a non-antimicrobial soap anddry hands and forearms completely. After application of the alcohol-basedproduct as recommended, allow hands and forearms to dry thoroughly beforedonning sterile gloves. Healthcare workers should also solicit information frommanufacturers regarding any effects that hand lotions, creams, or alcohol-basedhand antiseptics may have on the persistent effects of antimicrobial soaps beingused in the institution.