Back to Basics: Choosing the Appropriate Surface Disinfectant
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antibiotics
Review
Back to Basics: Choosing the Appropriate Surface Disinfectant
Angelica Artasensi , Sarah Mazzotta and Laura Fumagalli *
Dipartimento di Scienze Farmaceutiche, Università degli Studi di Milano, Via L. Mangiagalli 25, 20133 Milano,
Italy; angelica.artasensi@unimi.it (A.A.); sarah.mazzotta@unimi.it (S.M.)
* Correspondence: laura.fumagalli@unimi.it; Tel.: +39-0250319303
Abstract: From viruses to bacteria, our lives are filled with exposure to germs. In built environments,
exposure to infectious microorganisms and their byproducts is clearly linked to human health. In the
last year, public health emergency surrounding the COVID-19 pandemic stressed the importance of
having good biosafety measures and practices. To prevent infection from spreading and to maintain
the barrier, disinfection and hygiene habits are crucial, especially when the microorganism can persist
and survive on surfaces. Contaminated surfaces are called fomites and on them, microorganisms
can survive even for months. As a consequence, fomites serve as a second reservoir and transfer
pathogens between hosts. The knowledge of microorganisms, type of surface, and antimicrobial agent
is fundamental to develop the best approach to sanitize fomites and to obtain good disinfection levels.
Hence, this review has the purpose to briefly describe the organisms, the kind of risk associated
with them, and the main classes of antimicrobials for surfaces, to help choose the right approach to
prevent exposure to pathogens.
Keywords: antimicrobial; disinfectant; surface disinfection; fomite; surface contamination; microor-
ganisms
Citation: Artasensi, A.; Mazzotta, S.;
Fumagalli, L. Back to Basics:
Choosing the Appropriate Surface 1. Introduction
Disinfectant. Antibiotics 2021, 10, 613. In built environment, especially considering an indoor lifestyle, touching objects
https://doi.org/10.3390/ or surfaces which surround us is integral to everyday life. Such objects or surfaces if
antibiotics10060613
contaminated are called fomites and, in the 21st century, their role in disease transfer is
higher than ever in human history. Indeed, most microorganisms found in the indoor
Academic Editor: Gregory Caputo
environment are inactive, dormant, or dead and either show no impact on human health
or are even beneficial. Nevertheless, fomites can become contaminated by pathogenic
Received: 4 May 2021
organisms which have a variety of negative health consequences. In fact, microorganisms
Accepted: 17 May 2021
can survive even for many months and multiply on surfaces or objects [1], leading to the
Published: 21 May 2021
development of secondary reservoirs. As a consequence fomites can serve as a mechanism
for transfer between hosts, just think to doorknobs, elevator buttons, handrails, phones,
Publisher’s Note: MDPI stays neutral
with regard to jurisdictional claims in
keyboards, writing implement, etc., that are touched by a person that afterward will handle
published maps and institutional affil-
other objects (Figure 1).
iations.
Furthermore, experimental data show that touching a fomite carries approximately
the same risk for the acquisition of a lot of microorganisms (i.e., Methicillin-Resistant
Staphylococcus aureus—MRSA, Vancomycin-Resistant Enterococcus—VRE, and Clostridium
difficile) on hands as touching an infected patient [2–5]. Consequently, preventing trans-
mission of pathogens with disinfection procedures must be carried out not only in the
Copyright: © 2021 by the authors.
high-risk sectors, like laboratories, operating rooms, intensive care units, or food-handling
Licensee MDPI, Basel, Switzerland.
settings but also for hygienic behavior in everyday life on floors and on all the surfaces
This article is an open access article
distributed under the terms and
that frequently are touched with hands.
conditions of the Creative Commons
Therefore, environmental disinfection, hygiene habits, and the consequent mainte-
Attribution (CC BY) license (https:// nance of barriers are crucial in preventing infection from spreading. To develop effective
creativecommons.org/licenses/by/ policies and regulations to minimize the risk of transmission is strictly necessary to evalu-
4.0/). ate which organisms are present on the fomites. Furthermore, the choice of the effective
Antibiotics 2021, 10, 613. https://doi.org/10.3390/antibiotics10060613 https://www.mdpi.com/journal/antibiotics
to evaluate which organisms are present on the fomites. Furthermore, the choice of the
effective antimicrobial agent is also based on the risk assessment of the microorganisms
Antibiotics 2021, 10, 613 and the type of fomites 2 of 29
Public health emergency surrounding the COVID-19 pandemic, stressed the
importance of having good biosafety measures and practices, as never before. On these
bases, this review has the purpose to briefly describe the organisms, the kind of risk
antimicrobial agent is also based on the risk assessment of the microorganisms and the
associated with them, and the major characteristic of the main classes of antimicrobials for
type of fomites
surfaces to help in choosing the right approach to prevent exposure to pathogens.
Figure 1. Generic transmission route.
Figure 1. Generic transmission route.
2. Most Common
Public health Microorganisms on Fomites
emergency surrounding and Associated
the COVID-19 Risks stressed the impor-
pandemic,
tanceThe primary
of having goalbiosafety
good of disinfecting
measures procedures is the
and practices, as inactivation
never before.ofOn organisms
these bases,on
fomites. Generally,
this review microorganisms
has the purpose belong
to briefly to athe
describe diverse group the
organisms, suchkind
as bacteria, viral, and
of risk associated
protozoan
with species
them, and [6]. characteristic
the major These biologicalof theagents are widely
main classes found inforthe
of antimicrobials natural
surfaces to
environment and,the
help in choosing as right
a result, they can
approach to be foundexposure
prevent either in many work sectors or household
to pathogens.
contexts. The majority of these microorganisms are harmless; however, some of them or
2. Most
their Common may
metabolites Microorganisms on Fomites
cause diseases. and Associated
For example, Risks of norovirus that
the transmission
causesThe nonbacterial
primary goal ofgastroenteritis outbreaks
disinfecting procedures is theisinactivation
fomite-mediated
of organismsas onwell as
fomites.
Generally, microorganisms
coccidioidomycosis. belong tosome
Furthermore, a diverse group
of the such as
greatest bacteria,regarding
concerns viral, and protozoan
antibiotic-
species [6]. These biological agents are widely found in the natural environment and,
as a result, they can be found either in many work sectors or household contexts. The
majority of these microorganisms are harmless; however, some of them or their metabolites
may cause diseases. For example, the transmission of norovirus that causes nonbacterial
gastroenteritis outbreaks is fomite-mediated as well as coccidioidomycosis. Furthermore,
some of the greatest concerns regarding antibiotic-resistant bacteria transmission occur
Antibiotics 2021, 10, x FOR PEER REVIEW 3 of 32
Antibiotics 2021, 10, 613 3 of 29
resistant bacteria transmission occur via fomite as reported by Julian et al. [7] for
Staphylococcus pseudintermedius. Therefore, the knowledge of these organisms and their
survival
via fomite isas fundamental to choose
reported by Julian et al. the right
[7] for antimicrobial
Staphylococcus agents and implementing
pseudintermedius. Therefore,
effective
the tactics.
knowledge of these organisms and their survival is fundamental to choose the right
antimicrobial agents and implementing effective tactics.
2.1. Bacteria
2.1. Bacteria
Bacteria are single-celled organisms (0.3–1.5 µm) with independent life and
Bacteria cycle.
replication are single-celled organisms
Bacterial cells (0.3–1.5 µm)
are generally with independent
surrounded life and replication
by two concentric protective
cycle.
layers:Bacterial
an innercells
cellare generally surrounded
membrane and an outerbycell twowall
concentric
[8]. Theprotective layers:membrane
cytoplasmatic an inner
cell
shares a similar structure to the eukaryote’s one, but there are no sterols. Here, similar
membrane and an outer cell wall [8]. The cytoplasmatic membrane shares a proteins
structure
involvedtointhe theeukaryote’s one, but can
energy production therebeare no sterols.
found Here,
like some proteins involved
respiratory in theas
chain protein
energy production can be found like some respiratory chain protein as
well as photosynthetic protein in photosynthetic bacteria that lack chloroplast. Among thewell as photosyn-
thetic protein
proteins that in photosynthetic
constitute bacteria
the cell wall, the that
mainlack
onechloroplast. Among
is peptidoglycan the proteins
(PGN), also knownthatas
constitute the cell wall, the main one is peptidoglycan (PGN), also known
murein, which provides rigidity to the structure and counteracts the osmotic pressure of as murein, which
provides rigidityPGN
the cytoplasm. to theisstructure and counteracts
characterized by a glucidic thebackbone
osmotic pressure of the units
of alternating cytoplasm.
of two
PGN is characterized by a glucidic backbone of alternating units
azotated carbohydrates, namely N-acetylglucosamine (GlcNAc) and N-acetylmuramic of two azotated carbo-
hydrates, namelyEach
acid (MurNAc). N-acetylglucosamine (GlcNAc)
MurNAc is cross-linked to aand N-acetylmuramic
short amino acid chain, acid (MurNAc).
which can vary
Each MurNAc is cross-linked to a short amino acid chain, which can
with different bacterial species [9]. The differences in structural characterization vary with differentof
bacterial species define
peptidoglycan [9]. Thetwodifferences in structural
morphological characterization
categories: Gram-positiveof peptidoglycan define
and Gram-negative
two morphological categories: Gram-positive and Gram-negative bacteria (Figure 2).
bacteria (Figure 2).
Figure2.2.Gram-negative
Figure Gram-negativeversus
versusGram-positive
Gram-positivecell
cellwalls.
walls.
InInGram-positive
Gram-positivebacteria,
bacteria,peptidoglycans
peptidoglycansmake makeupupabout
about20%
20%ofofthethecell
cellwall
walldry
dry
weight;
weight; while in Gram-negative bacteria the thicker peptidoglycan layer containsabout
while in Gram-negative bacteria the thicker peptidoglycan layer contains about
10%
10%ofofthe
thecell
cellwall
walldrydryweight
weight[10].
[10].Furthermore,
Furthermore,Gram-positive
Gram-positivecell
cellwall
wallhas
hasa asignificant
significant
amount (up to 50%) of teichoic and teichuronic acid, which are involved
amount (up to 50%) of teichoic and teichuronic acid, which are involved in pathogenesisin pathogenesis
and
andplay
playkey
keyroles
rolesininantibiotic
antibioticresistance
resistance[11].
[11].
Certain
Certain bacteria may even haveaathird
bacteria may even have thirdoutermost
outermostprotective
protectivelayer
layercalled
calledaacapsule.
capsule.
Whip-like
Whip-like extensions often cover the surfaces of bacteria—long ones calledflagella
extensions often cover the surfaces of bacteria—long ones called flagellaand
and
short ones called pili—to become motile and seek out nutrients [12]. An alternative resource
short ones called pili—to become motile and seek out nutrients [12]. An alternative
exploited by some bacteria is the formation of endospores that are dormant and highly
resource exploited by some bacteria is the formation of endospores that are dormant and
resistant cells able to preserve the genetic material. This ruse helps the bacteria to survive
highly resistant cells able to preserve the genetic material. This ruse helps the bacteria to
even without nutrients or under extreme stress [13].
survive even without nutrients or under extreme stress [13].
Among endospore-producing bacteria, the most common are the Bacillus and
Among endospore-producing bacteria, the most common are the Bacillus and
Clostridium genera [14]. Table 1 reports several endospore-forming bacteria and their
Clostridium genera [14]. Table 1 reports several endospore-forming bacteria and their
relative clinical manifestations.
relative clinical manifestations.
otics 2021, 10, x FOR PEER REVIEW 4 of 32
Antibiotics 2021, 10, 613 4 of 29
Table 1. Common endospore-producing bacteria and their clinical manifestations.
Table 1. Common endospore-producing bacteria and their clinical manifestations.
Bacterial Species Clinical Manifestation
B. anthracis Bacterial Species anthrax Clinical Manifestation
B. cereus B. anthracis foodborne illness anthrax
B. subtilis B. cereus not pathogen foodborne illness
B. subtilis not pathogen
C. botulinum botulism
C. botulinum botulism
C. perfringens C. perfringens gas gangrene gas gangrene
C. tetani C. tetani tetanus tetanus
Another bacteria’s survival mechanism is the formation of biofilm: clusters of
Another bacteria’s survival mechanism is the formation of biofilm: clusters of bacteria
bacteria that are attached to a surface and/or to each other. During biofilm development,
that are attached to a surface and/or to each other. During biofilm development, bacteria
bacteria secrete extracellular
secrete extracellularpolymeric
polymeric substances
substances (EPS)
(EPS)which
whichare crucial to
are crucial tothe
theproduction of
production ofan an extracellular
extracellular matrix [15]. This network maintains cohesion between
matrix [15]. This network maintains cohesion between cells and the cells
and the surface and and
surface protects the the
protects accumulation
accumulation of of
microorganisms
microorganisms against
against chemical,
chemical, biological,
biological, and mechanical stressors. In this complex arrangement
and mechanical stressors. In this complex arrangement of cells, there of cells, thereareare
interstitial void
interstitial void spaces in which water flows so nutrients and oxygen
spaces in which water flows so nutrients and oxygen diffuse [16]. As biofilmdiffuse [16]. As protects from
biofilm protects from
harsh harsh conditions
conditions and resistance
and resistance towardstowards antibiotics,
antibiotics, it represents
it represents a seriousa global health
serious globalconcern.
health Furthermore,
concern. Furthermore, biofilm is involved in persistent
biofilm is involved in persistent chronic infections chronic
[17,18] and may
infections [17,18] and may
potentially potentially
contribute contribute
to their to their[19].
pathogenesis pathogenesis
In addition,[19].
someIn addition,
bacteria can produce a
some bacteriapolysaccharide
can produce aexocellular
polysaccharideslime exocellular slime which
(the glycocalyx), (the glycocalyx), which
adheres to compromised tissue
adheres to compromised tissue or the surfaces of biomaterials [20]. In fact, the glycocalyx
or the surfaces of biomaterials [20]. In fact, the glycocalyx is a fundamental factor in the
is a fundamental factor inofthe
persistence persistence
infection linkedof to
infection linked to
the prosthetic the prosthetic device.
device.
2.2. Virus 2.2. Virus
Viruses are
Viruses are subcellular subcellular
organisms organisms
with with submicroscopic
submicroscopic dimensions (nm).dimensions (nm). Their core
Their core
has either DNA (deoxyribonucleic acid) or RNA (ribonucleic
has either DNA (deoxyribonucleic acid) or RNA (ribonucleic acid) as genetic material. The acid) as genetic material.
core is coveredThebycore is covered
a protein by a called
coat [21], proteinthecoat [21], whose
capsid, called the
rolecapsid, whoseitrole
is to protect fromis to protect it
from degradation.
degradation. Furthermore, Furthermore,
the protein coat allowsthethe
protein
virus coat allows
to attach to athe virus receptor
specific to attach to a specific
receptor
of the host cell. In fact,ofviruses
the host arecell. In fact,
obligate viruses are
intracellular obligate[22],
parasites intracellular parasites
so they need host [22], so they
need host ribosomes
ribosomes to synthesize to synthesize
viral proteins. viral proteins.
Capsid proteins Capsid
are codified byproteins
the viralare codified by the viral
genome,
genome,
whose short length whose
entails short length
a limited numberentails a limited
of proteins withnumber offunction.
a specific proteins with a specific function.
This leads
to a capsid constituted by repetitive units of one or a few proteins combined in a combined
This leads to a capsid constituted by repetitive units of one or a few proteins
in a continuous
continuous structure [23], which structure
can have [23], which can
a helicoidal orhave a helicoidal
geometric symmetry.or geometric
The former symmetry. The
is characterized by a helicoidal distribution around the nucleic acid while the latter by while
former is characterized by a helicoidal distribution around the nucleic acid a the latter
polyhedral orbya aspherical
polyhedral or a spherical
shape. Besides theseshape. Besides
styles, theseviruses
a few styles, have
a fewaviruses
complex have a complex
architecture like poxviruses, geminiviruses, and many bacteriophages [24] (Figure 3). (Figure 3).
architecture like poxviruses, geminiviruses, and many bacteriophages [24]
Figure 3. Types of viruses architecture.
Figure 3. Types of viruses architecture.
Antibiotics 2021, 10, x FOR PEER REVIEW
Antibiotics 2021, 10, 613 5 of 29
Furthermore, some viruses show a further shell, called envelope, constituted
proteins and lipids. The envelope shields the virus from the immune system’s d
and,Furthermore,
in addition, facilitates
some the fusion
viruses show a furtherwith
shell,the host
called cell membrane
envelope, constituted[23].
by viral
proteins and lipids. The envelope shields the virus from the immune system’s detection
and, in addition, facilitates the fusion with the host cell membrane [23].
2.3. Fungi
2.3. Fungi
Fungi are a large group of eukaryotic organisms, mono or pluricellular, t
Fungiyeast
include are aand
largemolds.
group ofAseukaryotic organisms,
these organisms mono
have or pluricellular,
a rigid that also
cell wall (rich in chitin an
include yeast and molds. As these organisms have a rigid cell wall (rich in
polysaccharides, especially glucans as depicted in Figure 4) [25], they feed the chitin and other
polysaccharides, especially glucans as depicted in Figure 4) [25], they feed themselves
secreting
secreting digestive
digestive enzymes
enzymes and byand by absorbing
absorbing organic
organic matter frommatter from the environme
the environment: thus,
theyareare
they called
called heterotrophic
heterotrophic organisms.organisms.
Some fungi canSome fungi
live by can live
decomposing deadbyorganic
decomposin
organic
matter matterwhile
(saprobic) (saprobic) while
others are others
a parasite are a parasite
of organisms, of organisms,
even fungi, even fungi,
or have developed
complex symbionts as in lichens and mycorrhizae [26].
developed complex symbionts as in lichens and mycorrhizae [26].
Figure4.4.Fungal
Figure Fungalwall.
wall.
2.4. Microbiological Risk Assessment
2.4. According
Microbiological RiskofAssessment
to the Code Practice to the Safety, Health and Welfare at Work (Biological
Agents) Regulation to
According 2020the
[27]Code
the biological agents can
of Practice to be
theclassified
Safety,into four risk
Health groups,
and Welfare a
reported in Table 2. The classification takes into account:
(Biological Agents) Regulation 2020 [27] the biological agents can be classified i
•
risk Virulence—Ability
groups, reported of the microorganism to penetrate and multiplicate inside the host
in Table 2. The classification takes into account:
organism;
Virulence—Ability of
• Pathogenicity—Severity ofthe
thedisease
microorganism to penetrate and multiplicate in
that may result;
• host organism;
Transmissibility—Capability of the microorganism to be transmitted from one organ-
ism Pathogenicity—Severity
to another; of the disease that may result;
• Treatment—Availability, if
Transmissibility—Capability any, of effective
of theprophylaxis or therapy. to be transmitted fr
microorganism
Disinfection
organism to policies should be also based on risk assessment to control cross-
another;
contamination while reducing the risk caused by exposure to infectious agents. The
Treatment—Availability, if any, of effective prophylaxis or therapy.
evaluation of the surface’s risks and type together with the nature of the pathogen agent(s)
should lead to the use of an appropriate and effective antimicrobial agent. Such approaches
must be learned by everyone since their implementation in the routine measure improves
both cleaning performance and infection prevention [28].
However, as far as possible, the number of antimicrobials to be used should be limited
not only for healthy and economic reasons but also to reduce environmental pollution. Not
least, the discharge of waste biocides into the environment may promote the development
of both biocide and antibiotic resistance [29].
Antibiotics 2021, 10, x FOR PEER REVIEW 6 of 32
Table 2. Classification of biological agents.
Antibiotics 2021, 10, 613 6 of 29
Risk classification Description Examples Heading
Category 1 Pathogen with a low probability of Nonpathogenic strains
developing diseases in the humanof biological
Table 2. Classification of agents.
Escherichia
organism
Risk Classification Description Examples Heading
Category 2 Pathogen that may cause Measles virus,
Category 1 Pathogen with ain
pathology low probability
humans and ofbe a Nonpathogenic strains of
Salmonella, Legionella
developing diseases in the human Escherichia
potential hazard for workers; it’s
organism
unlikely that can be spread in the
Category 2 Pathogen that may cause pathology in Measles virus, Salmonella,
community; usually, there are
humans and be a potential hazard for Legionella
workers; it’s effective treatments
unlikely that can be spread
Category 3 Pathogen
in the community;thatusually,
may causetheresevere
are HIV, Bacillus anthracis,
effective treatments
illness in humans and be a serious HBV, HCV,
Category 3 hazard
Pathogen thatfor
may workers; the biological
cause severe Mycobacterium
illness in HIV, Bacillus anthracis, HBV,
humansagent and bemay
a serious hazard
spread in the for HCV, Mycobacterium
tuberculosis
workers; the biological agent may spread tuberculosis
community, but usually effective SARS-CoV-2
in the community, but usually effective SARS-CoV-2
treatments
treatments are are available
available
Category 4
Category 4
Pathogen that may cause severe
Pathogen that may cause severe illness in
Ebola virus, Lassa
Ebola virus, Lassa virus,
illness in humans and
humans and may be a serious hazard for may be a virus, Smallpox
Smallpox virus.virus.
workers;serious hazard for
the biological workers;
agent can spreadthe Nonpathogenic
Nonpathogenic strains
strains of
in thebiological
community, and usually,
agent can spread the Escherichia
thereinare Measles
of Escherichia virus,
Measles
no effective treatments available. Salmonella, Legionella
community, and usually, there are virus, Salmonella,
Pathogens with a low probability of
no effective
developing diseasestreatments available.
in human organisms. Legionella
Pathogens
Pathogens with
that may a low
cause probability
pathology in
humans and be a potential
of developing diseaseshazard for
in human
workers; it is unlikely that they can be
organisms. Pathogens that may
spread in the community; usually, there
cause arepathology in humans and be
effective treatments
a potential hazard for workers; it is
unlikely that they can be spread in
3. Factors That Affect the Activity of Antimicrobials
the community; usually, there are
The activity
effective of the antimicrobial agents depends on several factors, some of which are
treatments
intrinsic qualities of the organism, others derived from the chemicals and external physical
environment. More specifically
Disinfection need to
policies should bebealso
listed:
based on risk assessment to control cross-
• contamination
Number and type whileofreducing
microorganism No caused
the risk disinfectant can effectively
by exposure act on allagents.
to infectious microor-The
ganism classes.
evaluation So properrisks
of the surface’s choice
andoftype
chemical germicides
together with theisnature
fundamental. Furthermore,
of the pathogen agent(s)
some microbes
should lead to the can use
persist
of on
an surfaces showing
appropriate andresistance
effective to these products:
antimicrobial for ex-
agent. Such
approaches must be learned by everyone since their implementation in the from
ample, the production of endospores or biofilm matrix protects the pathogens routine
environmental
measure improves influences [13,30].
both cleaning performance and infection prevention [28].
• Type and concentration
However, as far as of the antimicrobial
possible, the number Afterof choosing the proper
antimicrobials to bedisinfectant,
used should the be
concentration
limited not onlyofforthehealthy
active ingredient
and economicis a key factor: but
reasons the influence of changing
also to reduce in the
environmental
concentration
pollution. of thethe
Not least, active(s) can be
discharge of measured experimentally,
waste biocides with the determination
into the environment may promote
of the kinetics of inactivation. Moreover, the knowledge
the development of both biocide and antibiotic resistance [29]. of the effect of dilution or
concentration on the activity of a sanitizing agent provides some valuable information
3. that could
Factors lead
that to athe
Affect reduction
Activityinof
the exposure time. Furthermore, microbicidal con-
Antimicrobials
centration is also a central concept in the microbial resistance field and it is especially
The activity of the antimicrobial agents depends on several factors, some of which
important nowadays with increasing knowledge and restrictions on the environmental
are intrinsic qualities of the organism, others derived from the chemicals and external
discharges of potentially harmful chemicals [31].
physical environment. More specifically need to be listed:
• pH of the solution The pH of the solution can affect the efficacy of the disinfection in
Number and type of microorganism
two ways: a change in the agent itself and a change in the interactions between the
No disinfectant can effectively act on all microorganism classes. So proper choice of
microbicide and the microbial cell. For example, several microbicides are effective
chemical germicides is fundamental. Furthermore, some microbes can persist on
in their unionized form (Table 3). Thus, the pH level would affect their degree of
surfaces showing
dissociation and would resistance
decreaseto these
their products:
overall forInexample,
activity. the production
contrast, other molecules of
are more effective in their ionized form. Besides these considerations, it should also
be kept in mind that any alteration of the pH level could affect the compound’s
stability. As a matter of fact, disinfectant products in the sanitary field are formulated
to guarantee, at a certain level of pH, maximum germicidal efficacy.
Antibiotics 2021, 10, 613 7 of 29
Table 3. Effects of pH level on antimicrobial activity.
Activity as
Classes of Disinfectants Mechanisms
Environmental pH Increases
Increase in the degree of dissociation of the
Phenols and organic acids
molecules
Decreased activity Undissociated hypochlorous acid is the most
Hypochlorites
fast-acting species
At low pH, iodine, the most powerful
Iodine
antimicrobial species, is the dominating one
Increase in the degree of ionization of
Quaternary ammonium compounds
Increased activity bacterial surface groups leading to an
(QACs)
increase in binding
• Formulation The formulation of a disinfectant deeply affects its activity. Several excip-
ients, such as solvents, surfactants, thickeners, chelating agents, colors, and fragrances,
can be found in these products; they can interact with the microorganisms or with the
active itself and ultimately affect the activity of the formulated product. Most of the
information on the effect of different excipients on the activity of disinfectants are not
available, since they are often trade secrets.
• Length of exposure The microbicidal activity of chemicals usually increases with the
rise of contact time. However, there is not a direct correlation between contact time
and microbicidal activity, maybe due to other factors. Contact times for disinfectants
are specific for each material and manufacturer. Therefore, all recommendations for
use of disinfectants should follow manufacturers’ specifications that must be reported
on the label.
• Temperature Temperature can be an important parameter that influences the pathogen’s
survival. High temperature can impact vital proteins and enzymes, as well as the
genome. Moreover, high temperature can boost and speed up the germicidal activity
of many chemicals resulting in reduced time and improved efficacy. As a drawback,
high temperature can accelerate the evaporation of the chemicals and also degrade
them. Particular care is needed in using and in stocking such chemicals in tropical
regions, where their shelf-life may be reduced because of high room temperature;
• Type of surfaces and precleaning process The location of microorganisms must be
considered as well: to sanitize an instrument with multiple pieces or joints and
channels is more difficult than a flat surface. Only surfaces that directly contact
the germicide will be sanitized. Indeed, the presence of dirt is the principal reason
for disinfection failure, since it could interact with the microbicide, reducing its
availability or interact with the microorganisms, giving protection. Moreover, material
characteristics of the surface may influence the survival of microorganisms as well:
for example, porous surfaces are more difficult to clean and, consequently, to disinfect.
Pretreatment of surfaces, especially when visibly soiled, is fundamental to ensure or
improve the microbicidal efficacy of the disinfection procedure.
Besides the activity that is influenced by the factors listed upon, ideally, an antimicro-
bial agent should: (1) have a wide spectrum against microorganisms; (2) be rapid in its
action; (3) be compatible with many materials; (4) be safe for humans and the environment.
4. Most Common Antimicrobial Classes
At present, there are numerous substances to be used on surfaces that are claimed
as antimicrobial agents and they are formulated alone or in combination. The most com-
mon disinfectants can be roughly divided as halogens, alcohols, quaternary ammonium
compounds (QACs), peroxigens, ozone, and UV. Generally, these antimicrobials damage a
specific part of the microorganism as reported in Figure 5.
Antibiotics 2021,
Antibiotics 10, x613
2021, 10, FOR PEER REVIEW 98of
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29
Figure 5. Mechanisms of biocide actions on microorganisms.
Figure 5. Mechanisms of biocide actions on microorganisms.
4.1. Halogens
4.1. Halogens
4.1.1. Chlorine Compounds
4.1.1. Historically,
Chlorine Compounds the most widely used antimicrobial agents belonging to halogens are
Historically,
chlorine and chlorine the most widely
releasing used antimicrobial agents belonging to halogens are
compounds.
chlorine and chlorine releasing compounds.
Since elemental chlorine gas (Cl2 ) is hazardous it must be banned either from work-
placesSince elemental environment
or household chlorine gasand (Clsubstituted
2) is hazardous it must be banned
by chlorine-releasing agents.either from
workplaces
The most or household
commonlyenvironment and substituted
used chlorine-releasing agentby is chlorine-releasing
sodium hypochlorite agents.
(NaOCl),
The most
universally commonly
known used which
as bleach, chlorine-releasing
is characterizedagentby is high
sodium hypochlorite
microbicidal (NaOCl),
efficacy, low
universally known asand
toxicity to humans, bleach,
low which
cost, but is characterized by high microbicidal
suffers the disadvantages of beingefficacy,
irritant low
and
toxicity
corrosive. to Nevertheless,
humans, and ceramics,low cost, methylacrylate,
but suffers the or disadvantages
cement are not of sensitive
being irritant and
to bleach.
More specifically, sodium hypochlorite is potentially bactericidal,
corrosive. Nevertheless, ceramics, methylacrylate, or cement are not sensitive to bleach. virucidal, fungicidal,
mycobactericidal,
More specifically, sporicidal. Hence it plays
sodium hypochlorite an important
is potentially role in the surface
bactericidal, virucidal, disinfection
fungicidal,of
healthcare facilities
mycobactericidal, and medical
sporicidal. Hence equipment.
it plays an important role in the surface disinfection
The concentration
of healthcare of sodium
facilities and medicalhypochlorite
equipment. sold for domestic purposes is around 5–6%,
withThea pH around 11 and
concentration it is irritant;
of sodium while in higher
hypochlorite sold forconcentration,
domestic purposes 10–15%, with a pH
is around 5-
around 13, it burns and it is corrosive. According to the Laboratory
6%, with a pH around 11 and it is irritant; while in higher concentration, 10-15%, with a biosafety manual [32]
published
pH around by 13, the World
it burns andHealth Organisation
it is corrosive. (WHO):
According general all-purpose
“ALaboratory
to the biosafetylaboratory
manual
disinfectant should have a concentration of 1 g/L available chlorine.
[32] published by the World Health Organisation (WHO): “A general all-purpose laboratoryA stronger solution, containing
5 g/L available
disinfectant shouldchlorine,
have aisconcentration
recommendedoffor dealing
1 g/L with chlorine.
available biohazardous spillagesolution,
A stronger and in the presence
containing
5ofg/L
large amounts
available of organic
chlorine, matter. Sodium
is recommended hypochlorite
for dealing solutions, as spillage
with biohazardous domesticand bleach,
in thecontain 50
presence
g/L available chlorine and should therefore be diluted 1:50 or 1:10 to obtain
of large amounts of organic matter. Sodium hypochlorite solutions, as domestic bleach, contain 50 final concentrations of
1 g/L and 5 g/L, respectively. [ . . . ] Surfaces can be decontaminated using
g/L available chlorine and should therefore be diluted 1:50 or 1:10 to obtain final concentrations ofa solution of sodium
1hypochlorite
g/L and 5 g/L, (NaOCl); a solution
respectively. […]containing
Surfaces can 1 g/L
be available chlorineusing
decontaminated may be suitable for
a solution general
of sodium
environmental sanitation, but stronger solutions (5 g/L) are recommended
hypochlorite (NaOCl); a solution containing 1 g/L available chlorine may be suitable for general when dealing with
high-risk situations.”
environmental sanitation, but stronger solutions (5 g/L) are recommended when dealing with high-
Once sodium hypochlorite dissolves in water (Equations (1)–(3)) the two compounds
risk situations.”
that Once
causesodium
disinfection via oxidation
hypochlorite dissolves areingenerated, namely (1)–(3))
water (Equations hypochlorite
the twoion (OCl− ), a
compounds
weak
that base,disinfection
cause and its corresponding
via oxidation acid,arehypochlorous acid (HOCl),
generated, namely whose ion
hypochlorite percentage
(OCl−), is
a
determined by water’s pH and which is the most active between the two [33,34]. In fact,
weak base, and its corresponding acid, hypochlorous acid (HOCl), whose percentage is
Antibiotics 2021, 10, x FOR PEER REVIEW 10 of 32
determined by water’s pH and which is the most active between the two [33,34]. In fact,
Antibiotics 2021, 10, 613 hypochlorous acid, due to no electronic charge, better penetrate the microorganism 9cell
of 29
wall or any protective layer and effectively kills them by oxidating the side chains of
proteins’ amino acids [35,36].
hypochlorous acid, due to no electronic + ⇆
charge, + penetrate the microorganism cell wall
better (1)
or any protective layer and effectively kills them by oxidating the side chains of proteins’
amino acids [35,36]. + + ⇆ + (2)
OCl − + H2 O HOCl + OH − (1)
2 + → + 2 + (3)
HOCl + H + + Cl − Cl2 + H2 O (2)
It is also common to express the concentration of chlorine compounds in terms of
2HOCl + OCl − → ClO− + 2Cl − + H + (3)
available chlorine or free available chlorine (FAC).3 The term FAC refers to the mixture of
oxidizing
It is chlorine
also commonforms to that have athe
express chlorine atom in of
concentration thechlorine
0 or −1 oxidation
compounds state
in and
terms areof
not combined with ammonia or organic nitrogen.
available chlorine or free available chlorine (FAC). The term FAC refers to the mixture of
Sodium
oxidizing hypochlorite
chlorine forms is characterized
that have a chlorine by high
atominstability, −1 oxidation
in the 0 ortherefore the FACstatevalue is
and are
not
notsocombined
significant: 0.75
with grams of
ammonia oractivated chlorine evaporate per day. This happens not
organic nitrogen.
only when
Sodium sodium hypochlorite
hypochlorite gets heatedby
is characterized up,high
but instability,
also when therefore
gets in touch withvalue
the FAC acids,is
not so significant:
sunlight, 0.75 grams
specific metals, toxic and of activated
corrosivechlorine evaporate
gases, included per day.
chlorine This
itself happens not
[37,38].
onlySodium
when sodium hypochlorite
hypochlorite solutiongets is anheated up, butweak
inflammable also when
base and getsthese
in touch with acids,
characteristics
sunlight, specific metals, toxic and corrosive gases, included chlorine
must be considered during its use and storage. Because of these reasons, formulation and itself [37,38].
Sodium
conditions for hypochlorite
the application solution
should is an inflammable
minimize weak base
the formation ofand these characteristics
by-products and even
must be considered
chloramines [39]. Theduring
overallits use and storage.
stoichiometry Because of is
of degradation these
shownreasons, formulation
in Equation 3. and
conditions for the application
Thus disinfection’s efficacyshould
of chlorineminimize the agents
releasing formation of by-products
depends and even
on the water’s pH
chloramines
and FAC. Chlorine [39]. The overall stoichiometry
disinfection against vegetative of degradation
bacteria, isfungi,
shown and in yeast,
Equation (3). as
as well
Thus disinfection’s efficacy of chlorine releasing agents
fungal conidia and viruses, is preferable at alkaline NaOCl solutions; although depends on the water’s pHthe
and
FAC. Chlorine disinfection against vegetative bacteria,
germicidal efficacy is even greater when pH value is around 5.5 and 8 [39,40]. fungi, and yeast, as well as fungal
conidia and viruses,
Furthermore, Kuroiwais et preferable
al. [41] at alkaline
proved thatNaOCl solutions;
adjusting the pH although
aroundthe5 germicidal
by weak
acidification with acetic acid, resulted in a shortened killing time of all the B.Kuroiwa
efficacy is even greater when pH value is around 5.5 and 8 [39,40]. Furthermore, subtilis
et al. [41]
JCM1465 proved
spores by that adjusting
one-third. On thethecontrary,
pH around this5preparation
by weak acidification
killed all ofwith acetic acid,
the non-spore-
resultedbacteria
forming in a shortened
within killing
30 secondstime asof all the B.as
quickly subtilis
NaClO JCM1465
solution spores
withoutby one-third.
acidification.On the
The importance of the pH level is shown in Figure 6. At a pH of 7, the concentration ofas
contrary, this preparation killed all of the non-spore-forming bacteria within 30 seconds
quickly as NaClO
hypochlorous acid issolution
80%, whilewithoutwhen acidification.The
the pH value isimportance
around 8, the of the pH level is drops
concentration shown
in Figure
to 20%. 6. At a pH of 7, the concentration of hypochlorous acid is 80%, while when the
pH value is around 8, the concentration drops to 20%.
Figure 6. Active chlorine species concentration at different pH values [42].
Figure 6. Active chlorine species concentration at different pH values [42].
The pH value of the solution is fundamental either for the bactericidal activity or
for the shelf life: at 25–35 ◦ C, neutralized-NaOCl solutions (pH 7) expires in a few hours,
generated NaOCl (gNaOCl) solutions (produced by electrolysis of a salt (NaCl solution,
pH 9) last 6 days, while stabilized NaOCl solutions (pH 9–11) persist more than 30 days [43].
The pH value of the solution is fundamental either for the bactericidal activity or for
Antibiotics 2021, 10, 613
the shelf life: at 25–35 °C, neutralized-NaOCl solutions (pH 7) expires in a few 10 hours,
of 29
generated NaOCl (gNaOCl) solutions (produced by electrolysis of a salt (NaCl solution,
pH 9) last 6 days, while stabilized NaOCl solutions (pH 9–11) persist more than 30 days
[43].
Sodium
Sodiumhypochlorite
hypochlorite isiswidely
widelyused,
used,notnotonly
onlyas
asaasurface
surfaceantimicrobial
antimicrobialbutbutalso
alsoinin
water
watertreatment,
treatment,water
waterdisinfection,
disinfection,and andbleaching
bleachingin inthe
thetextile
textileindustry.
industry.Furthermore,
Furthermore,itit
can
canbe
beused
usedto toavoid
avoidcrustaceans
crustaceansand andalgae
algaeformation
formationinincooling
coolingtowers.
towers.
As
As an alternative, calcium hypoclorite (Ca(OCl)2 2) also known as
an alternative, calcium hypoclorite (Ca(OCl) ) also known asHTH
HTH(high
(hightest
test
hypochlorite)
hypochlorite) can be used as well. HTH is sold in granular form that, once in solution,
can be used as well. HTH is sold in granular form that, once in solution,
achieves
achievesaapH pHof of9–11
9–11and
andititisisas
asstable
stableas
asNaOCl
NaOCl[43].
[43].
Another
Anotherchlorine
chlorinereleasing
releasingagent
agentthat
thathas
hasbeen
beenexplored
exploredas asan
analternative
alternativeto
tosodium,
sodium,
or
orcalcium,
calcium,hypochlorite
hypochloriteisissodium
sodiumdichloroisocyanurate
dichloroisocyanurate(NaDCC).
(NaDCC).This Thiscompound
compoundisisthe
the
sodium salt of a chlorinated hydroxytriazine (Figure
sodium salt of a chlorinated hydroxytriazine (Figure 7). 7).
Figure7.7.Structure
Figure Structureof
ofsodium
sodiumdichloroisocyanurate
dichloroisocyanurate(NaDCC).
(NaDCC).
Thisdisinfectant
This disinfectantisisavailable
availableas asaastable
stablepowder
powderthat thatproduces
producessolutions
solutionsthat thathave
haveaa
pH level of around 6 and expire within hours [43]. These solutions
pH level of around 6 and expire within hours [43]. These solutions are more susceptible are more susceptible
toinactivation
to inactivation by by organic
organicmatter
matter than
than NaOCl
NaOCl [44–46].
[44–46]. NaDCC
NaDCC is is often
often used
used as as aa broad-
broad-
spectrumdisinfectant
spectrum disinfectantsincesinceitithas
hasbeen
beenreported
reportedto togenerally
generallyachieve
achievesimilar
similardisinfection
disinfection
activities to
activities to chlorine, while
whileresults
resultstotobebeless lesscorrosive.
corrosive. OnOnstainless steel,
stainless Bloomfield
steel, Bloomfieldet al.
[47]
et al. reported lower
[47] reported ME ME
lower (microbiocidal
(microbiocidal effect) values
effect) following
values followinga 5-minutes exposureto
a 5-min exposure to
250 ppm NaDCC
250 NaDCCcompared
comparedtoto NaOCl
NaOCl at the samesame
at the concentration
concentrationagainst S. aureus
against (2.4 vs.
S. aureus
(2.4
4.9 vs. 4.9 tolog
to 3.6 and2%0.3 porklog
meat) after 30
reductions seconds exposure
(without/with 2% pork to 0.2%
meat)(w/v)afterNaOCl.
30 secondsInterestingly,
exposure the entry(w/v)
to 0.2% containing
NaOCl.
hypochlorite
Interestingly,as theanentry
antibacterial
containing agent and anionactive
hypochlorite tensides as cleaning
as an antibacterial agent andcompounds
anionactive
were
tensides as cleaning compounds were considered to be much more efficient (3.8 and2%
considered to be much more efficient (3.8 and 2.2 log reductions, without/with 2.2
pork meat) thanwithout/with
log reductions, the hypochlorite 2% pork disinfectant,
meat) than probably due to the
the hypochlorite inactivation
disinfectant, of the
probably
NaOCl
due to by
thethe organic matter.
inactivation of the NaOCl by the organic matter.
To
To be effectiveagainst
be effective againstbacteria
bacteriaand andspores,
spores,an anadequate
adequateconcentration
concentrationof ofHOCl
HOClisis
required; in Table 4 are reported the recommended dilutions of
required; in Table 4 are reported the recommended dilutions of each chlorine releasing each chlorine releasing
compound
compound mentioned
mentioned until until now
now totosignificantly
significantlyreducereducethe therisk
riskofoftransmission.
transmission. The The
surface
surface conditions, the main advantages, and drawbacks have also beenconsidered.
conditions, the main advantages, and drawbacks have also been considered.
4.1.2. Iodine Compounds
Although less reactive than chlorine, iodine solution has a broad spectrum of antimi-
crobial activity against both gram-negative and gram-positive bacteria, fungi, protozoa,
and even bacterial spores [12], while it is not so effective as virucidal [50]. Many investiga-
tions identified elemental iodine I2 and hypoiodous acid (HIO) as the two most powerful
antimicrobials agents among the several iodine species.
I2 + H2 O H IO + I − + H + (4)
H IO IO− + H + (5)
3H IO + 3OH − IO3− + 2I − + 3H2 O (6)Antibiotics 2021, 10, 613 11 of 29
Table 4. Recommended dilutions of commonly used chlorine releasing compounds.
Use Condition
Chlorine Type Advantages Disadvantages
Clean Condition Dirty Condition
Can be local (stabilized form) Shorter shelf life
Sodium hypochlorite
Can be on-side Difficult to ship
solution (5% available 20 mL/L 100 mL/L
(no stabilized form) Low stability
chlorine)
Does not clog pipes (no stabilized form)
High-test hypochlorite Easy to ship
1.4 g/L 7.0 g/L Explosive
(70% available chlorine) Long shelf life
Sodium
Easy to ship
dichloroisocyanurate
1.7 g/L 8.5 g/L Long shelf life Smell
powder (60% available
Does not clog pipes
chlorine)
Sodium
Easy to ship
dichloroisocyanurate
1 tablet per L 4 tablets per L Long shelf life Smell
tablets (1.5 g available
Does not clog pipes
chlorine per tablet)
The dissociation constant of hypoiodous acid is 4.5 × 10−13 and it reveals that the
formation of hypoiodite ion (IO− ) in an aqueous solution is insignificant. The percentages
of the species (see Equations (4)–(6)) are directly related to the pH level of the solution and,
to a much lesser extent, to the temperature.
Figure 8 shows I2 hydrolysis data at different pH values and it is clear that the highest
concentrations of the antimicrobial species are present in the acid range. In fact, when the
solution is alkaline, several iodine species that have no apparent antimicrobial activity can
Antibiotics 2021, 10, x FOR PEER REVIEW 13 of 32
also be generated. Iodate formation could not be a problem if the pH value stays below 8
and the contact time of disinfection is accomplished in the first 30 min.
Figure
Figure8.8.pH-dependent
pH-dependentspeciation
speciationof
ofiodine
iodine[51].
[51].
Historically solutionsorortinctures
Historically iodine solutions tinctures have
have beenbeen primarily
primarily used used by health
by health profes-
professionals as antiseptics
sionals as antiseptics on skinon or skin or Unfortunately,
tissue. tissue. Unfortunately,
aqueous aqueous
solutionssolutions
are generallyare
generally unstable
unstable so so a combination
a combination of iodine andof iodine and a solubilizing
a solubilizing agenthas
agent or carrier or carrier has been
been formulated.
formulated. These combinations,
These combinations, called
called iodophor, iodophor,
have have
been used been
both as used both asand
antiseptics antiseptics and
disinfectants,
retaining the retaining
disinfectants, germicidalthe efficacy of iodine
germicidal but being
efficacy morebut
of iodine stable andmore
being relatively
stable free of
and
toxicity and
relatively freeirritancy [39].and
of toxicity They have been
irritancy [39].developed
They havetobeen
slowly release iodine
developed (I2 ) from
to slowly the
release
iodine (I2) from the complex, which can be a cationic surfactant, non-ionic, polyoxymer,
or polyvinylpyrrolidone [52]
The most known and widely used iodophor is povidone-iodine, Figure 9. Regarding
this complex, Block et al. observed 3.14, 3.49, 3.47, and 3.78 log reduction, after 1.5 min forFigure 8. pH-dependent speciation of iodine [51].
Historically iodine solutions or tinctures have been primarily used by health
professionals as antiseptics on skin or tissue. Unfortunately, aqueous solutions are
Antibiotics 2021, 10, 613 generally unstable so a combination of iodine and a solubilizing agent or carrier has been
12 of 29
formulated. These combinations, called iodophor, have been used both as antiseptics and
disinfectants, retaining the germicidal efficacy of iodine but being more stable and
relatively free of toxicity and irritancy [39]. They have been developed to slowly release
complex,
iodine (I2which
) from can
the be a cationic
complex, surfactant,
which can be anon-ionic, polyoxymer,
cationic surfactant, or polyvinylpyrroli-
non-ionic, polyoxymer,
done [52].
or polyvinylpyrrolidone [52]
The
Themost
mostknown
knownand andwidely
widelyused
usediodophor
iodophorisispovidone-iodine,
povidone-iodine,Figure
Figure9.9.Regarding
Regarding
this complex, Block et al. observed 3.14, 3.49, 3.47, and 3.78 log reduction,
this complex, Block et al. observed 3.14, 3.49, 3.47, and 3.78 log reduction, after after
1.51.5
minminfor
for VRE, E. faecalis, and methicillin-resistant and methicillin-sensitive S. aureus, respec-
VRE, E. faecalis, and methicillin-resistant and methicillin-sensitive S. aureus, respectively
tively
[53]. [53].
I3
N O H O N N O
C C CH 2 CH C CH
H2 H H2
n m
Figure9.9.Structure
Figure Structureof
ofpovidone-iodine
povidone-iodinecomplex.
complex.
Surfactantiodophor,
Surfactant iodophor,whenwhenused,
used,may
mayaddaddaafurther
furtherdetergency
detergencyactivity,
activity,even
eventhough
though
iodine
iodine isischemically
chemicallylesslessreactive
reactivethan
thanchlorine.
chlorine. Moreover,
Moreover, surfactant
surfactant iodophor
iodophor isislessless
affected
affectedby bythe
thepresence
presenceofoforganic
organicmatter
matterthan
thanchlorine.
chlorine.
An
Aniodophor,
iodophor, when
when used at 25at
used ppm25 (parts
ppm per million
(parts per ofmillion
available
of iodine), is considered
available iodine), is
to act as a sanitizer,
considered to act ashowever, when
a sanitizer, the same
however, product
when is applied
the same at 75
product is ppm fallsatinto
applied the
75 ppm
disinfectant
falls into thecategory.
disinfectant category.
Antibiotics 2021, 10, x FOR PEER REVIEW 14 of 32
After
Afteritsitsrelease,
release,iodine
iodinecan
canquickly
quicklypenetrate
penetratethe
thecell
cellwall
wallofofaamicroorganism
microorganismand and
oxidize
oxidizethiol
thiolgroups
groupsleading
leadingtotodisruption
disruptionofofproteins
proteinsand
andnucleic
nucleicacids
acidsstructures
structures[39].
[39].
4.2.
4.2.Alcohols
Alcohols
4.2.1.
4.2.1.Alifatic
AlifaticAlcohols
Alcohols
Among
Amongthe theseveral aliphatic
several alcohols
aliphatic thatthat
alcohols exhibit microbicidal
exhibit properties
microbicidal ethyl alco-
properties ethyl
hol (ethanol), isopropyl alcohol (isopropanol, propan-2-ol), and n-propanol are
alcohol (ethanol), isopropyl alcohol (isopropanol, propan-2-ol), and n-propanol theare
most
the
commonly used (Figure
most commonly 10). 10).
used (Figure
Figure10.
Figure 10.Antimicrobial
Antimicrobialalcohols.
alcohols.
Thesedisinfectants
These disinfectantsare arerapid
rapidbactericidal
bactericidalrather
ratherthan
thanbacteriostatic
bacteriostaticagainst
againstvegetative
vegetative
bacteria,included
bacteria, includedmycobacteria
mycobacteriabut but have
have nono effect
effect onon spores.
spores. TheThe bactericidal
bactericidal properties
properties of
of ethanol
ethanol were
were examined
examined against
against several
several microorganisms
microorganisms fordifferent
for differentranges
rangesofoftime
time[54]:
[54]:
P.P.aeruginosa,
aeruginosa, Serratia
Serratia marcescens, E. coli,coli, and Salmonellatyphy
andSalmonella typhywere
werekilled
killedin in
1010 s by all
seconds by
concentrations
all concentrations of ethanol from
of ethanol from40%40%to 100%
to 100% (30%
(30%forfor
thetheE.coli entry).S.S.aureus
entry).
E.coli aureusandand
Streptococcus
Streptococcuspyogenes
pyogenes were
wereslightly more
slightly resistant,
more beingbeing
resistant, killed killed
in 10 s with
in 10concentrations
seconds with
of 60%–95%. Isopropyl
concentrations alcoholIsopropyl
of 60%–95%. resulted slightly
alcohol more bactericidal
resulted than ethyl
slightly more alcoholthan
bactericidal for
E. coli and
ethyl S. aureus
alcohol for E.[55]. Furthermore,
coli and this category
S. aureus [55]. of biocides
Furthermore, shows limited
this category fungicidal
of biocides shows
and virucidal
limited activity
fungicidal especially
and virucidal onactivity
lipophilic viruses such
especially as herpesviruses
on lipophilic virus, influenza virus,
such as herpes
and hepatitis B and C viruses [56,57]. Literature data demonstrate that
virus, influenza virus, and hepatitis B and C viruses [56,57]. Literature data demonstrate isopropyl alcohol
shows its antimicrobial
that isopropyl alcohol activity
shows its against lipid viruses
antimicrobial but itagainst
activity is not active
lipid against
virusesthebutnonlipid
it is not
enteroviruses
active against[58] the nonlipid enteroviruses [58]
These
Thesealcohols
alcohols exert their
exert antimicrobial
their antimicrobialactivity by causing
activity proteinprotein
by causing denaturation [59,60].
denaturation
Water plays an important role in the formulation of alcoholic disinfectants
[59,60]. Water plays an important role in the formulation of alcoholic disinfectants because, in its
because, in its absence, proteins are not readily denatured by alcohol. Therefore a 70%
solution of alcohol is a much more effective sanitizer than the pure (99%) product [61], but
when the concentration drops below 50% there is no practical value [62]. Concentration
can be expressed both by weight/weight percentage (%w/w) and, most frequently, byAntibiotics 2021, 10, 613 13 of 29
absence, proteins are not readily denatured by alcohol. Therefore a 70% solution of alcohol
is a much more effective sanitizer than the pure (99%) product [61], but when the concen-
tration drops below 50% there is no practical value [62]. Concentration can be expressed
both by weight/weight percentage (%w/w) and, most frequently, by volume/volume per-
centage (%v/v). This value is important since it is linked to the evaporation rate: a higher
concentration of alcohol evaporates quickly. The evaporation speed could be an issue if a
longer contact time is requested, but the addition of surfactants [63], or combination with
alkali, mineral acids, and hydrogen peroxide could overcome this problem [12,24].
Alcohols are fast-acting, easy to use but are not free from limitations that are due to
poor detergent properties, toxicity, and, of course, their flammability, which is a big concern.
The minimum temperature at which vapors above a volatile combustible substance ignite
in air when exposed to flame defines the flashpoint. The higher the concentration, the
lower the flashpoint. For example, the flashpoints of 70% ethyl and 70% isopropyl alcohol
are 20.5 ◦ C and 21.0 ◦ C, respectively, while the flashpoint of 30% ethyl alcohol is 29 ◦ C [64].
Moreover, even if alcoholic disinfectants are neither corrosive nor staining, they could
damage some instruments, by swelling or hardening rubber.
4.2.2. Aromatic Alcohols
Besides aliphatic alcohols, also aromatic ones exhibit antimicrobial properties being
effective in sanitization and disinfection, even in the presence of biological fluids. Phe-
nols are the reference standard for the Rideal–Walker (RW) and Chick–Martin tests for
disinfectant evaluation [65].
Phenol (C6 H5 OH) is an organic compound that consists of a benzene ring bearing a
single hydroxy substituent. It appears as a white crystalline solid, which is partially water-
soluble (1 g/15 mL water) [66] and it has a pKa value of 10, which means it is classified as
a weak acid.
Phenol exerts its antimicrobial activity against vegetative bacteria, both Gram-positive
and negative, fungi and viruses but it is not so effective as sporicidal and against
acid-fast bacteria.
The biological activity is related to the undissociated molecule, which induces progres-
sive leakage of essential metabolites, including the release of K+ [67], leading to membrane
damage and consequentially cell lysis, while acting like a protoplasmic poison causing
coagulation of the cytoplasm [68].
Phenol is the parent compound but the chemical structure can be modified by replacing
one of the hydrogens on the aromatic ring with a different functional group (halogen, alkyl,
phenyl, benzyl, etc.). Figure 11 represents several microbicidal phenols.
The structure-activity relationship in the phenol series was investigated by Suter [69].
Regarding the results, it is interesting to notice that the microbiocidal activity increases
in derivatives with alkyl chain in the para position, constituted by a maximum of six
carbon atoms, since for longer chain the activity drops probably due to the decrease of
water solubility. Nitrophenols were evaluated as well; unfortunately, the toxicity increased
towards both bacteria and humans and there is also a trend to be inactivated by organic
matter. Finally, bisphenolic compounds show activity if they are connected by a methyl
linker, sulfur, or oxygen atom, and even if they are directly linked. Augmentation of the
efficacy can also be achieved by halogen substitutions.
Among all the derivatives, o-phenylphenol and 2-benzyl-4-chlorophenol are widely
used as healthcare disinfectants.
As disclosed by published reports, commonly used phenolic compounds show, at their
use dilution, antimicrobial efficacy against bacteria, fungi, viruses, including HIV [70–73].
However, literature reports also that the phenolic disinfectants ‘Stericol’ and ‘Lysol’ show a
limited effect on Coxsackie B4, Enterovirus 11, and Poliovirus [74].You can also read
