Karin M.
Stoffels – Schülke & Mayr, Germany
Antimicrobial stabilisation
of cosmetic products
The regulatory framework for preservatives
varies in different regions. According to
the EU Cosmetics Directive 76/768/EEC
preservatives are substances ‘for the primary
purpose of inhibiting the development
of microorganisms’ in cosmetic products.
These substances are listed in Annex VI
of the EU Cosmetics Directive.1 Japan lists
preservatives in Annex III of the Japanese
Standard of Cosmetics.2 Under the Federal
Food, Drug, and Cosmetic Act in the US,
cosmetic ingredients, with the exception
of colour additives, ‘do not require FDA
approval before they go on the market.
Companies who market cosmetics have
the legal responsibility to ensure the safety
of their products’.3
In addition to legal requirements,
the options for microbial stabilisation
are restricted by marketing demands.
The movement to preservative-free or
natural products and restrictions on certain
actives, i.e. formaldehyde-releasers,
isothiazolinones, etc., are increasing.
The desire to avoid cosmetic
preservatives extends to all categories
of traditional preservatives, leading to
a limited number of accepted actives.
Parabens, for example, are some of the
most commonly used preservatives due to
their excellent efficacy and low sensitising
potential.4,5 Public discussion has caused
some manufacturers to avoid using these
materials in new formulations.
The claim ‘paraben-free’ on personal
care products first appeared in 2005.
According to market research company
Mintel, in 2011 this claim was used for
9% of total beauty and personal care
launches worldwide; mainly in North
America, Europe, and Japan. In contrast,
the ‘preservative-free’ claim is still of minor
importance. Less than one per cent of
new cosmetic products were launched
OH
HO
Figure 1: Phenethyl alcohol. Figure 2: Ethylhexylglycerin.
PRESERVATIVES
ABSTRACT
The microbial safety of cosmetic products
is an important consideration. It is integral
to ensuring the stability of the products for
the entire shelf- life and to preventing
damage to health of the consumers.
Currently, the number of accepted
substances for microbial stabilisation is
limited due to legal restrictions or public
discussion. Hurdle technology and the
intelligent combination of multifunctional
additives help to design microbiologically
www.stockfreeimages.com
stable products. The synergistic blend of
phenethyl alcohol and ethylhexylglycerin is
an effective stabilisation system, while
consisting of 90% nature-identical
material.
worldwide with this claim; mainly in Japan combined with some natural ingredients.
(3% of total launches).6 It is debatable The ‘truly natural’ criteria often include
whether these claims meet the consumer higher portions of natural ingredients and
needs or increase their uncertainty of the mainly avoid less desirable ingredients
safety of cosmetic products. such as silicones, ethoxylates, chemical
In addition, the trend to natural and UV filters, and synthetic preservatives.7
sustainable products is ongoing. According However, products including high
to Kline & Company, the global natural percentages of natural components
personal care segment in 2010 is about are more susceptible to microbial
USD 23 bn. This is a small segment in contamination, either because the raw
the personal care market (~USD 300 bn). material contains a high initial bio-burden
However, the sales growth of 15% from or because most natural materials are an
2009 to 2010 and the expected annual excellent medium for microbial growth.
sales growth of 12% through 2015 are Due to these obstacles, formulators are
remarkable.7,8 interested in finding novel ways to keep
There is still no universal definition of cosmetic products microbiologically stable.
‘natural’ within the personal care market.
Looking into the ingredients of natural Technologies in use
products, Kline differs in ‘truly natural’ and One formulation technique to achieve self-
‘natural-inspired’ products and states that preserving formulations or to reduce the
76% of global sales of natural products need for preservatives is known as hurdle
are in the group of ‘natural-inspired’. technology. This technique has been used
The majority of ingredients in natural- in the food industry since the 1970s.9
inspired products are synthetic. These are Hurdle technology describes intelligent
formulation using different preservation
factors. Good manufacturing practices,
CH3
OH appropriate packaging, careful choice of
the form of emulsion, low water activity
O CH3 and low or high pH values can be used to
control microbial growth in the absence
of traditional preservatives.10
Additionally, chelating agents can be
November 2012 P E R S O N A L C A R E 95
 PRESERVATIVES

Escheria coli ATCC No. 11229 Candida albicans ATCC No. 10231
107 107

106 106

105 105

104

cfu/mL
cfu/mL

104

103 103

102 102
■ 0.05% ethylhexylglycerin ■ 0.05% ethylhexylglycerin
10 ■ 0.45% phenethyl alcohol 10 ■ 0.45% phenethyl alcohol
■ 0.50% sensiva PA 20 ■ 0.50% sensiva PA 20
0 0
0 3 6 24 48 72 168 0 3 6 24 48 72 168
Hours Hours
Figure 3: Germ count reduction tests in oil-in-water emulsion with E. coli and C. albicans.

used to increase the efficacy of
antimicrobial substances. Chelating agents
remove metal ions from the cell membrane
of the microbes forming complexes. The
lack of metal ions weakens the cell
membrane and allows a better penetration
of the preservatives.11
A common chelating agent providing this
effect is ethylenediaminetetraacetic acid
(EDTA). Due to widespread use and poor
biodegradability, EDTA has emerged as a
persistent organic pollutant in the
environment.12 Alternatives to EDTA are
readily biodegradable chelating agents; e.g.
the sodium salts of glutamate diacetate,
iminodisuccinate, and ethylenediamine
disuccinate. These complexing agents
provide a comparable or potentially better
boosting effect of the antimicrobial
stabilisation system phenoxyethanol/
ethylhexylglycerin than does EDTA.
Adjusting the pH of the formulation with
citric acid instead of hydrochloric acid
increases this boosting effect.13
However, formulators are often limited
and cannot implement all factors of hurdle
technology, driven by marketing requirements
for the cosmetic product. Cosmetic
additives providing antimicrobial capacity or
a boosting effect can close the gap.
Multifunctional additives
Multifunctional additives are molecules with
more than one beneficial effect to the
formulation or the skin; e.g. glycols, glycerol
ethers, fragrance ingredients, and essential
oils. Some of these widely-used
multifunctional additives also display a
certain antimicrobial efficacy. The synergistic
combination of multifunctional additives can
be used to achieve the antimicrobial
stabilisation of cosmetic formulations
without using traditional preservatives.
Glycols
Glycols are broadly used humectants in
cosmetic products. Their antimicrobial
efficacy increases corresponding to their
chain lengths, whereas the water solubility
decreases with longer chain lengths. This
leads to a lower antimicrobial effect in the
formulation for linear glycols with more than
eight C-atoms. A secondary effect is that
glycols lower the water activity, which helps
maintain the microbial stability of cosmetic
preparations.
Phenethyl alcohol
Phenethyl alcohol occurs widely in nature
and can be found in many essential oils;
including rose, carnation and hyacinth.
It is used as a fragrance component and
has a mild rose odour. It exhibits effective
inhibitory action on microbes, especially
Gram-negative bacteria.
The mechanism of action of phenethyl
alcohol is at the level of the cell
membrane, causing a breakdown of the
cellular permeability barriers. This alteration
of membranes is assumed to lead to
disruption of DNA and protein synthesis.14
Ethylhexylglycerin
Ethylhexylglycerin is a multifunctional
cosmetic ingredient with excellent
deodorising and skin care properties. It has
a limited activity against spoilage-causing
bacteria. However, if combined with
alcohols, antimicrobial stabilisers like
glycols or preservatives, ethylhexylglycerin
acts as a booster, enhancing their
antimicrobial activity.15
The chemical structure must be
considered to explain the mode of action of
this boosting effect. Due to its surfactant-
like structure, ethylhexylglycerin has the
capability to significantly reduce the surface
tension of water. This improves the contact
of the antimicrobial actives with the cell
membranes resulting in better penetration
and therefore a higher efficacy.15

Staphylococcus aureus ATCC No. 6538 Aspergillus niger ATCC No. 16404
107 107

106 106

105 105
cfu/mL

104
cfu/mL

104

103 103

102 102
■ 0.05% ethylhexylglycerin ■ 0.05% ethylhexylglycerin
10 ■ 0.45% phenethyl alcohol 10 ■ 0.45% phenethyl alcohol
■ 0.50% sensiva PA 20 ■ 0.50% sensiva PA 20
0 0
0 3 6 24 48 72 168 0 3 6 24 48 72 168
Hours Hours

Figure 4: Germ count reduction tests in oil-in-water emulsion with S. aureus and A. niger.

96 P E R S O N A L C A R E November 2012
 PRESERVATIVES

Solution for antimicrobial

stabilisation
The combination of ethylhexylglycerin with the
fragrance ingredient phenethyl alcohol has been
investigated with regard to its antimicrobial
properties and to a possible boosting effect of
ethylhexyglycerin on phenethyl alcohol.

Test material
All described tests have been done with nature-
identical phenethyl alcohol. Nature-identical
materials, synthetic versions of the same
chemistries found in nature, offer a way to
harness the best of nature without relying on
the availability or quality of the materials that
exist in nature.
Sensiva SC 50 was used as single ingredient
ethylhexylglycerin. The test combination sensiva
PA 20 is a mixture of both ingredients in the
ratio of 9:1.

Methods
Germ count reduction test
The purpose of the germ count reduction test is
Biotech innovations
to determine exposure times for antimicrobial
substances. Oil-in-water emulsions are
WITH A TOUCH
inoculated in the laboratory. The titre of the
suspension is approx. 109 cfu/mL for bacteria,
OF NATURE
108 cfu/mL for yeasts and 107 cfu/mL for
moulds. Nutrient media are CSA (tryptone-soya-
agar) for bacteria and SA (sabouraud-dextrose-
agar) for yeast and mould. The plates are
incubated for 48 hours at 37˚C for all tested
germs except the mould Aspergillus niger
which is incubated for 48 hours at 25˚C.

Challenge test (Schülke Koko test)

A mixture of bacteria, yeast and moulds are
inoculated 6 times (once a week) into the test
material, with the goal of keeping the test
material germ free for this period. The inoculum
contains pathogenic microorganisms as germs
which are well known for product spoilage.
All species have to be cultivated separately and
mixed directly before the addition, to ensure a
constant composition and germ count of the
inoculum. Its germ count is approximately VITIPURETM, VELSAN® and ZENVIVOTM
108-9 cfu/mL, which means a germ count of A unique range of biopolymers for personal care to fulfill
about 106 cfu/mL in the sample. Nutrient media consumers desire for purity and effectiveness. These
are CSA (tryptone-soya-agar) for bacteria and personal care ingredients originate from natural sources
SA (sabouraud-dextrose-agar) for yeast and and contribute to consumers’ well-being and well-feeling.
moulds. Moisturizing, anti-aging and protecting effects are
combined with an extraordinary gentle feel for skin and hair.
Results
Figures 3 and 4 show germ count reduction
tests in oil-in-water emulsion for the bacteria
Staphylococcus aureus and Escherichia coli,
CLARIANT INTERNATIONAL LTD
as well as the mould Aspergillus niger and
Rothausstrasse 61
the yeast Candida albicans.
4132 Muttenz
The tests have been conducted with
Switzerland
phenethyl alcohol and ethylhexylglycerin
individually, and with a mixture in the ratio of
WWW.PERSONALCARE.CLARIANT.COM
9:1 (sensiva PA 20).
Phenethyl alcohol achieved a reduction
of the spoilage causing germs, whereas
ethylhexylglycerin in this small amount

November 2012 P E R S O N A L C A R E 97
 PRESERVATIVES

had no effect when used alone. The killing Table 1: Oil-in-water emulsion. Table 2: Water-in-oil emulsion.
rate of phenethyl alcohol was increased
significantly for all germs by pairing it with Phase Ingredients % (w/w) Phase Ingredients % (w/w)
ethylhexylglycerin at a ratio of 9:1. A Aqua ad 100 Aqua ad 100
Pure phenethyl alcohol (0.45%/0.9%)
Glycerin 85% 2.30 Glycerin 85% 0.90
reduced all germs to zero after 168 hours.
The synergistic blend achieved the reduction Sodium Hydroxide 0.20 Propylene Glycol 0.50
to zero after 48 hours for C. albicans and (45% aqueous solution) Sodium Hydroxide 0.50
S. aureus and after 6 hours for E. coli. (45% aqueous solution)
A faster decrease was even observed in Sodium Choloride 0.15
the persistent mould A. niger. B Paraffinum Liquidum 6.00
The synergistic effect of the blend has Dimethicone 5.00 B Cetyl PEG/PPG-10/1 0.80
also been shown in challenge tests Stearic Acid 4.00 Dimethicone
(Schülke Koko test) with two formulations
Cetyl Alcohol 3.00 Cera Alba 0.35
(Table 1-3). Based on experience, a
cosmetic product without growth of Glyceryl Stearae, 3.00 Hydrogenated Castor Oil 0.25
microorganisms after six inoculation PEG-30 Stearate Paraffinum Liquidum 7.30
cycles (six weeks) can be considered Octyl Palmitate 1.50 Carbomer 0.70
microbiologically stable for 30 months,
which is recommended for cosmetic
products.16 Conclusion interpretation), as well as sensitive
The combination of phenethyl alcohol The contamination of cosmetic products applications; e.g. baby care, oral and lip
with ethylhexylglycerin (sensiva PA 20) is a high risk. The concept of hurdle care. Due to the high portion (90%) of
stabilises the o/w cream well at a level technology is not always sufficient to nature-identical material in this blend,
of 1%. The w/o cream is well stabilised achieve self-preserving systems or not all it can be an option to stabilise nature-
at a level of 0.75%. The single ingredients necessary factors can be implemented. inspired cosmetics. PC

in the corresponding concentrations show The intelligent combination of

no efficacy in the Koko test (Table 1).
Ethylhexylglycerin was found to be an
excellent booster for the antimicrobial
capacity of phenethyl alcohol. The killing
rate in the germ count reduction tests
was accelerated. The boosting effect
was confirmed by the results of the
challenge tests. Depending on the
composition of the formulations,
0.75% and 1% of a blend of these two
components stabilised the emulsions
effectively.
Table 3: Challenge test (Schülke Koko test) with two formulations.
Test material
0 1
Oil-in-water cream, unpreserved a rBMY
+ 0.90% Phenetyl Alcohol a rBMY
+ 0.10% Ethylhexylglycerin a rBMY
+ 0.675% Phenetyl Alcohol a •M
+ 0.075% Ethylhexylglycerin
(0.75% sensiva PA 20)
+ 0.90% Phenetyl Alcohol a a
+ 0.10% Ethylhexylglycerin
(1% sensiva PA 20)
Water-in-oil cream, unpreserved a rBMY
+ 0.675% Phenetyl Alcohol a uB
+ 0.075% Ethylhexylglycerin a rBMY
+ 0.675% Phenetyl Alcohol a a
+ 0.075% Ethylhexylglycerin
(0.75% sensiva PA 20)
Key
a = Free of microbial growth
M = Moulds
• = Slight growth (<103 cfu/mL)
r = Massive growth (>105 cfu/mL)
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98 P E R S O N A L C A R E November 2012