COMPARISON OF BACTERIAL RECOVERY FROM PERSONAL CARE

PRODUCTS USING BIOCONTROL’S SIMPLATE® TOTAL PLATE COUNT

COLOR INDICATOR SYSTEM AND TRADITIONAL POUR-PLATE METHODS

COMPARISON OF BACTERIAL RECOVERY FROM PERSONAL CARE PRODUCTS USING
BIOCONTROL’S SIMPLATE® TOTAL PLATE COUNT COLOR INDICATOR SYSTEM AND
TRADITIONAL POUR-PLATE METHODS

Abstract

The SimPlate® Total Plate Count Color Indicator (TPC-CI) test is a unique system
for the detection and quantification of total aerobic microorganisms in cosmetics
and toiletries. It offers results after 24 - 28 hours providing a significant time
saving advantage over the current culture methods. The SimPlate’s Binary
Detection TechnologyTM (BDT) is comprised of a proprietary media formulation
in conjunction with a patented plating device. The SimPlate TPC-CI method was
compared to a conventional Letheen agar pour plate method (CTFA
Microbiological Guidelines) for quantifying total aerobic microorganisms in nine
different commercially available products, representing a variety of preservative
systems and cosmetic matrices, and four challenge organisms (Escherichia coli,
Pseudomonas aeruginosa, Pseudomonas cepacia, and Staphylococcus aureus).
The two methods demonstrated comparable recovery of all four organisms for
eight of the nine matrices and SimPlate demonstrated higher recovery of P.
aeruginosa from the ninth matrix.
The SimPlate TPC-CI method was found to be comparable to the CTFA reference
method. However, SimPlate results were available 24 hours sooner than pour
plate results providing a distinct advantage to manufacturers wishing to reduce
inventory holding costs and release product as quickly as possible.
 COMPARISON OF BACTERIAL RECOVERY FROM PERSONAL CARE PRODUCTS USING BIOCONTROL’S
SIMPLATE® TOTAL PLATE COUNT COLOR INDICATOR SYSTEM AND TRADITIONAL POUR-PLATE METHODS
SCOPE
To validate the efficacy of the SimPlate Total
Plate Count-Color Indicator (TPC-CI) system
for quantifying the total bacterial count of
selected personal care products as compared to
the Cosmetics Toiletries & Fragrances M-1
reference method “Determination of the
Microbial Content of Cosmetic Products”.
DESIGN OF STUDY
Overview
Nine products representing a variety of
preservatives systems were evaluated (Table
1). A sample from each product was
neutralized and artificially inoculated with 3
gram negative strains and 1 gram positive
strain. An aliquot from each of the inoculated
products was then analyzed via the SimPlate
method and the reference method. After the
required incubation time, bacterial populations
were enumerated and final counts were
converted to base 10 logarithms (log10) for data
comparison.
Product Neutralization and Initial Dilution
For the water-miscible products, 10g of sample
was added to 90ml of sterile Letheen broth
(Difco) creating a 1:10 dilution of the sample.
For water-immiscible products, 10g sample
was initially added to a dilution bottle
containing 10ml of Polysorbate 80. The
sample was dispersed in the Polysorbate 80 and
Letheen broth was added to bring the final
volume to 100ml creating a 1:10 dilution of the
sample.
Artificial Contamination of the Neutralized
Product
A total of 180 test samples representing 5
replicates of 9 products inoculated with 4
challenge microorganisms (Escherichia coli
2
ATCC 8739, Pseudomonas aeruginosa ATCC
9027, Pseudomonas cepacia ATCC 25416, and
Staphylococcus aureus ATCC 6538) were
analyzed. Each replicate received a low level
inoculum of organisms (one strain per tube).
Negative controls, consisting of 10g of product
in 90ml of Letheen broth (no challenge
organisms) were prepared for each product.
Positive controls consisted of separate tubes of
Letheen broth (no product) inoculated with the
challenge organisms, one strain per tube.
Test Methods
After product neutralization, the diluted test
samples, positive controls and negative
controls were analyzed with both the reference
and SimPlate methods.
Reference method: The method described in
CTFA M-1, section 4, Procedure for Aerobic
Plate Count was used as a guideline for the
reference method. One milliliter from the
artificially contaminated neutralized sample
was transferred to a sterile petri dish in
duplicate. Fifteen milliliters of molten Letheen
agar, tempered to 44-48°C, were added to each
plate. Plates were then gently swirled to
evenly distribute the sample and agar. All
plates were incubated inverted at 35°C for 48 ±
2 h. All plates were counted after the required
incubation and recorded.
SimPlate method: The same diluted sample
used for the reference method was also tested
by the SimPlate method. Testing was
conducted according to the manufacturer’s
instructions for use. One milliliter from the
artificially contaminated neutralized sample
was transferred to SimPlate device. The
samples were overlayed with 9 ml of
rehydrated SimPlate TPC-CI medium
containing lecithin and Tween 80. The devices
were gently swirled to distribute the
sample/medium mixture into all the wells.
Excess liquid was decanted into the collection
sponge. Plates were incubated upright at 35°C
 COMPARISON OF BACTERIAL RECOVERY FROM PERSONAL CARE PRODUCTS USING BIOCONTROL’S
SIMPLATE® TOTAL PLATE COUNT COLOR INDICATOR SYSTEM AND TRADITIONAL POUR-PLATE METHODS
for 24-28 h. The total bacterial count was controls. Target organisms were recovered
determined by converting the number of from all positive controls.
positive wells with the SimPlate Conversion
Table (Appendix 1). Conclusion

Data Analysis The data generated in this study indicate that

The final microorganisms per gram of product
generated from the SimPlate and reference
methods were converted to base 10 logarithms
(log10) for data comparison (Table 1).
RESULTS AND DISCUSSION
The equivalence of the SimPlate TPC-CI
method and the CTFA M-11 aerobic plate
count method at quantifying bacterial
populations in representative samples of
personal care products were compared. A total
of 180 test samples representing 5 replicates of
9 products inoculated with 4 challenge
microorganisms were analyzed in this study.
Table 1 summarizes the bacterial mean log
counts for both methods. Each of the nine
products was grouped into one of four
categories Sunscreens, Facial Masks, Lotions/
Creams, or Body Washes. A Correlation
Coefficient (r) and the Average Difference of
the Mean Logs were calculated for each these
four product groups (Table 2).
the SimPlate TPC-CI method is comparable to
the CTFA reference method for the
enumeration of total microbial counts in all
nine of the personal care products evaluated.
SimPlate TPC-CI represents a cost effective
alternative to the traditional pour plate method
for the detection and quantification of total
aerobic microorganisms in cosmetics &
toiletries. SimPlate offers the advantage
providing results in 24 - 28 hours, 1 day faster
than the reference method, which reduces
product holding time providing a financial
benefit to the manufacturer. The SimPlate
method also provides easy to read results,
eliminating the uncertainty of enumeration that
may be caused by over crowding, spreading,
and product particulate interference in the
traditional pour- plate-method.

The two methods demonstrated comparable

recovery of all organisms from eight of the
nine matrices tested and for three of the four
organisms from the ninth matrix. The
SimPlate method demonstrated greater
recovery of P. aeruginosa from the Smoothing
Shower Scrub (Table 1). There was good
correlation between methods for all four
product groups. The average difference of the
Mean Logs was less than 0.25 for all four
groups.

There were no positive SimPlate wells and no

growth on the Letheen agar for the negative

1
CTFA Microbiology Guidelines, 2000 The Cosmetic
Toiletry & Fragrance Association
3
 COMPARISON OF BACTERIAL RECOVERY FROM PERSONAL CARE PRODUCTS USING BIOCONTROL’S
SIMPLATE® TOTAL PLATE COUNT COLOR INDICATOR SYSTEM AND TRADITIONAL POUR-PLATE METHODS

Table 1. Comparative Results for Aerobic Plate Counts by the SimPlate Method and the
Letheen Agar Pour Plate Method

Product Type Target Organism Mean Log Counta Mean log

SimPlate Referenceb difference
Sunscreens
E. coli 1.60 1.52 0.08
P. aeruginosa 1.78 1.54 0.24
Sunscreen SPF 30
P. cepacia 1.04 0.90 0.15
S. aureus 2.02 1.95 0.07
Peel Off Masks
E. coli 1.58 1.42 0.15
Peel Off Mask P. aeruginosa 1.85 1.36 0.49
P. cepacia 1.18 1.00 0.18
S. aureus 2.03 1.95 0.09
Lotions/Creams
E. coli 1.67 1.61 0.07
P. aeruginosa 1.91 1.59 0.32
Body Luster
P. cepacia 1.28 1.16 0.12
S. aureus 2.02 1.93 0.09

E. coli 1.66 1.52 0.14

P. aeruginosa 1.67 1.25 0.42
Moisture Rich Body Lotion
P. cepacia 1.27 0.90 0.37
S. aureus 1.94 1.97 -0.03

E. coli 1.64 1.52 0.12

P. aeruginosa 1.74 1.30 0.44
Body Cream
P. cepacia 1.14 1.04 0.10
S. aureus 2.09 1.89 0.20

E. coli 1.65 1.52 0.13

P. aeruginosa 1.75 1.36 0.39
Night Cream
P. cepacia 1.33 1.14 0.19
S. aureus 1.98 1.98 0.00
Body Washes
E. coli 1.59 1.35 0.24
P. aeruginosa 1.92 1.58 0.34
Creamy Body Wash
P. cepacia 1.88 1.75 0.13
S. aureus 1.72 1.75 -0.03

E. coli 1.43 1.34 0.09

P. aeruginosa 1.82 1.07 0.75
Smoothing Shower Scrub
P. cepacia 1.74 1.52 0.22
S. aureus 1.76 1.57 0.18

E. coli 1.53 1.40 0.13

Aromatherapy Foaming P. aeruginosa 1.83 1.54 0.29
Bath P. cepacia 0.76 0.71 0.05
S. aureus 1.09 0.90 0.19
a
mean log aerobic bacteria/g for 5 independent replicates.
b
Letheen Agar pour plate based on CTFA M-1 method

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 COMPARISON OF BACTERIAL RECOVERY FROM PERSONAL CARE PRODUCTS USING BIOCONTROL’S
SIMPLATE® TOTAL PLATE COUNT COLOR INDICATOR SYSTEM AND TRADITIONAL POUR-PLATE METHODS

Table 2. Correlation of SimPlate and Letheen Agar Pour Plate Results by

Product Group
Product Group Correlation of Methods Average Difference of
(r) Mean Logs
Sun Screen 0.98 0.14
Facial Mask 0.87 0.21
Lotions / Creams 0.90 0.19
Body Washes 0.83 0.22

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Appendix 1 SimPlate Conversion Table