1770 International Journal of Food Science and Technology 2009, 44, 1770–1777

Original article
Thermal resistance of Alicyclobacillus acidoterrestris spores in
different heating media

Gulin Ceviz,1 Yahya Tulek2* & Ahmet Hilmi Con2

1 Ministry of Agricultural and Rural Affairs, Directorate of Control Laboratory, Denizli, Turkey
2 Food Engineering Department, Pamukkale University, Engineering Faculty, TR-20070 Kinikli, Denizli, Turkey
(Received 25 February 2009; Accepted in revised form 6 May 2009)

Summary The aim of this work was to study the influence of temperature (85, 90, 95 and 100 C), total soluble solids
(SS: 10 and 20Brix or % by weight of sucrose) and pH (3.5 and 4.0) on decimal reduction time (D-value) of
the Alicyclobacillus acidoterrestris strain DSM2498 spores in apple juice, orange juice and malt extract broth
(MEB). The effects of SS and pH on D-values and z-values in each media were insignificant (P > 0.05). In
apple juice, orange juice and MEB, z-values of A. acidoterrestris for pH 3.5 and pH 4.0 were 12.2 ± 1.3–
14.2 ± 3.2 C, 11.2 ± 0.3–9.4 ± 0.0 C and 11.9 ± 0.8–10.3 ± 0.4 C, respectively. z-values of apple
juice, orange juice and MEB samples with SS = 10Brix and SS = 20Brix were 14.1 ± 3.2–12.2 ± 1.3 C,
10.2 ± 0.7–10.5 ± 1.1 C and 11.3 ± 1.5–10.9 ± 0.2 C, respectively. However, D-values of all samples
were affected by temperature significantly (P < 0.01). Average D-values of apple juice, orange juice and
MEB were 101.2 ± 14.7, 34.4 ± 7.9, 20.3 ± 4.9 and 4.3 ± 1.3 min for 85, 90, 95 and 100 C. This study
demonstrated that A. acidoterrestris spores exhibited high resistance to thermal processing applications. pH
and SS of the media did not affect thermal resistance.
Keywords Alicyclobacillus acidoterrestris, apple juice, D-value, orange juice, thermal resistance, z-value.

been isolated and identified in several spoiled commer-

Introduction
Thermal processing is applied to store fruit juices for a
long time without spoiling and to eliminate pathogenic
micro-organisms. As fruit juices have acidic character-
istics (pH < 4.5), temperatures of 85–95 C interval are
sufficient for the pasteurisation process. As a result of
the thermal processing carried out in that specific
temperature interval, vegetative forms of bacteria are
inactivated. On the other hand, the bacterial spores,
which could still survive in the foodstuff after the
thermal processing, cannot germinate or grow as a result
of acidic conditions. However, Cerny et al. (1984)
reported an unusual spore-forming bacterium that
spoiled aseptically packaged apple juice in Germany.
This bacterium was subsequently named Bacillus acido-
terrestris (Deinhard et al., 1987). Later, Wisotzkey et al.
(1992) proposed a new genus, Alicyclobacillus, charac-
terised by x-alicyclic fatty acids as the major natural
membrane lipid component. It was re-identified as
Alicyclobacillus acidoterrestris.
Alicyclobacillus acidoterrestris is a thermoacidophilic,
non-pathogenic and spore-forming bacterium that has
*Correspondent: Fax: +90 258 2963262;
e-mail: ytulek@pau.edu.tr
cial pasteurised fruit juices, such as apple and orange
juices. Several authors have reported thermoacidophilic
bacteria from spoiling fruit juices. Yamazaki et al.
(1996) isolated and identified A. acidoterrestris in spoiled
acidic juices. Splittstoesser et al. (1994) isolated acidic
spore-forming Bacillus spores (VF strain) from a spoiled
apple juice, later identified as A. acidoterrestris by
Pontius et al. (1998).
This micro-organism is unique in that it contains x-
alicyclic fatty acids as the major membrane fatty acid
component in its cells (Chang & Kang, 2004), and is able
to grow in a pH range of 2.0 to 7.0 and a temperature
range of 20–70 C (Palop et al., 2000). The optimal
growth temperature for A. acidoterrestris is 42–53 C
depending on the strain. Pettipher et al. (1997) discov-
ered that A. acidoterrestris can grow in apple juices and
orange juices at 25 C, 35C and 44 C whereas it can
not grow below 4 C. According to the results of
Bahceci et al. (2003), the spore population after depec-
tinisation carried out at 50 C for 2 h was found to be
higher than that carried out at 25 C for 24 h because of
the thermophilic structure of Alicyclobacillus.
Alicyclobacillus acidoterrestris spores are highly resis-
tant to high temperatures. They can survive in pasteuri-
sation norms applied in the fruit juice industry. As a

doi:10.1111/j.1365-2621.2009.01995.x
 2009 The Authors. Journal compilation  2009 Institute of Food Science and Technology

result of the germination and growth of these spores
surviving in juices, taste and aroma defects can be
occured (Jensen, 2000). Therefore, this micro-organism
has been suggested as the target to be used in the design
of adequate pasteurisation processes of acidic food
products (Murakami et al., 1998; Silva et al., 1999; Silva
& Gibbs, 2001, 2004). Indeed, A. acidoterrestris spore
germination and growth to 106 cfu mL)1 under acidic
conditions has also been reported in orange juice stored
at 44 C for 24 h (Pettipher et al., 1997). Komitopoulou
et al. (1999) reported that A. acidoterrestris spores
germinate and grow in apple, orange and grapefruit
juices stored at 30 C. Pontius et al. (1998) showed that
A. acidoterrestris can survive for 2 min at 88–96 C
during the hot filling processes. A. acidoterrestris have
been determined to germinate and grow best under the
conditions of pH 4.5 and 43 C (Sinigaglia et al., 2003).
Some preservatives used in the preservation of food-
stuffs reduce the thermal resistance of micro-organisms
(Acar & Cemeroglu, 1998). As well as the studies
concerning the temperature variables, some other stud-
ies have been conducted on how A. acidoterrestris is
affected by several chemicals (Pettipher & Osmundson,
2000; Yamazaki et al., 2000; Lee et al., 2004). Lee et al.
(2004) investigated the reduction of A. acidoterrestris
spores in several culture media and apples with chlorine
dioxide. It was found that increasing the concentration
of chlorine dioxide and extending the application time
result in significant increases in the inactivation of
A. acidoterrestris spores.
The preservative (sorbic acid, benzoic acid or both)
use in fruit juices is reported to have inhibitive effects on
A. acidoterrestris vegetative cells and spores (Pettipher &
Osmundson, 2000). Nisin addition is determined to
reduce thermal resistances of A. acidoterrestris spores in
fruit juices and thus being effective on the inhibition of
germination and growth of these bacterial spores
(Yamazaki et al., 2000). Bevilacqua et al. (2008)
searched the antimicrobial effect of sodium benzoate
and some natural compounds like cinnamaldehyde,
eugenol and limonene against outgrowth of A. acidotr-
restris spores (c8 and c4). These researchers determined
that limonene compound had no effect on the inhibition
of the relevant spores. However, decreases in between
57% and 70% at spore outgrowth was observed for
samples stored 13 days at 44 C added with 100 ppm
cinnamaldehyde or sodium benzoate, whereas com-
pletely hindered spore outgrowth was established in the
presence of 500 ppm eugenol. Separately, it was con-
cluded that cinnamaldehyde was the efficient compound
at inhibition of outgrowth, and strain c8 was more
resistant compared with the other strain c4. Similar
researchers, in their other recent study (Bevilacqua
et al., 2009) determined the critical cinnamaldehyde
concentration for outgrowth inhibition of A. acidoter-
restris spores during the 3 and 10 days of storage at
 2009 The Authors. Journal compilation  2009 Institute of Food Science and Technology
Thermal resistance of A. acidoterrestris spores G. Ceviz et al. 1771
44 C, after applying heat treatment between 80 and
96 C. In the relevant study, decrease at critical
cinnamaldehyde concentration in accordance with pH
reduction at medium was reported.
The effects of temperature and high pressure on the
inactivation of A. acidoterrestris have been both
separately and jointly investigated. Alpas et al. (2003)
reported that there is a significant increase in the
inactivation of A. acidoterrestris cells in model system
(BAM broth), orange juice, apple juice and tomato
juice. Lee et al. (2002) and Buzrul et al. (2005) explained
that the processing temperature has obvious and signif-
icant effects on the pressure inactivation. For rais-
ing threat of A. acidoterrestris at fruit juice industry,
Walker & Phillips (2008) evaluated the applicable
isolation techniques and rapid identification methods,
spoilages created by these bacteria and the novel
applications for preventing spoilages in their extensive
review.
The thermal resistance characteristics of food-spoilage
micro-organisms are quite important determinants of
the design of the thermal processes used to preserve
various food products. As put forward by many
researches, A. acidoterrestris is an important source of
microbial spoiling in fruit juices. Only limited numbers
of studies have been carried out to determine the
thermal resistance characteristics of this bacterium.
More studies are required to be carried out especially
on the determination of thermal resistance characteris-
tics of various forms of this bacterium under various
media conditions. Accordingly, the aim of this study is
to determine the thermal resistance characteristics of
A. acidoterrestris spores under different pH, total
soluble solids (SS) and temperature values in real food
media such as apple juice and orange juice as well as
malt extract broth (MEB) model media.
Materials and methods
Material
Alicyclobacillus acidoterrestris strain DSM2498 (Ger-
man Collection of Micro-organisms and Cell Cultures,
Braunschweig, Germany) spores were used in the study.
The apple juice concentrate and orange juice concentrate
(SS: 69Brix) were obtained from KONFRUT Fruit
Juice Factory in Denizli ⁄ Turkey. Malt extract broth
(MEB; Merck, Darmstadt, Germany) as the model
medium and BAT agar (Merck, Darmstadt, Germany)
as the counting and sporulation medium were used.
Preparation of the spore suspension
The original A. acidoterrestris in slant agar was taken
from its culture with the help of a sterile loop and put in
the tube filled with sterile physiological water. BAT agar
International Journal of Food Science and Technology 2009
1772 Thermal resistance of A. acidoterrestris spores G. Ceviz et al.
was inoculated and spread with obtained culture
suspension, and incubated for 5 days at 45 C. The
culture obtained 5 days later was applied Gram staining
and spore staining for confirmed of sporulation and
purity. A. acidoterrestris growing in Petri plates was
scraped with distilled water and then put in sterile
centrifuge tubes. Centrifugation was applied for 15 min
on 5000 rpm in the refrigerated centrifuge (Hettic,
Universal 30 RF, Tuttlingen, Germany) adjusted to
4 C to eliminate A. acidoterrestris from the suspension.
A. acidoterrestris spores accumulated in the bottom of
the centrifuge tubes. After the elimination of the
liquid on top of the tubes, 2 mL of sterile distilled water
was added to the sediment and then it became homog-
enous by vortex. Washing and centrifuging processes
were repeated for five times under aseptic conditions.
After the last centrifuging, the pellet was added distilled
water and transferred to sterile glass tubes with screwed
lids after homogenised by using vortex (Sahbaz et al.,
1996).
The vegetative cells in the tubes containing about
108spores mL)1 were inactivated by keeping the tubes at
80 C for 10 min. Thus the pure spore culture was
achieved (Pettipher et al., 1997). The culture obtained
was stored in the refrigerator at 4 C until use. The
number of spores in 1 mL of the spore suspension was
determined at the each usage.
Experimental design
Apple juice, orange juice and MEB solutions were
prepared to ensure that they would have two different
SS (10 and 20Brix or % by weight of sucrose) and two
different pH (3.5 and 4.0) values. Thermal processing was
applied under four different temperatures (85, 90, 95 and
100 C) for each sample. Five different thermal process-
ing periods were used for each temperature in the light of
preliminary experiments and literature knowledge
(Table 1). The experiments were carried out as two
parallel and two replication. They were carried out on
completely randomised factorial design. Statistical anal-
ysis of the data was performed using the Minitab and
Mstad computer programs (Freed, 1991). When anova
revealed a significant effect (P < 0.01), data means were
compared by the least significant difference test (Freed,
1991).
Table 1 Heating temperatures and heating time
Temperature (°C) Heating time (min)
85 20 40 60 80
90 5 10 15 20
95 1⁄2 1 2 4 8
100 1⁄6 1⁄2 1 3⁄2
International Journal of Food Science and Technology 2009
Thermal processing media
Apple juice, orange juice and MEB were used as the
thermal processing media. Apple and orange juice
concentrates provided in aseptic packages (SS: 69Brix)
were diluted with distilled water whereas MEB was
added sucrose (Merck) to adjust to the expected Brix (SS
or % by weight of sucrose) values. The SS adjustments
were made at 20 C by using a refractometer (RFM 340
Model; Bellingham–Stanley Ltd., London, UK) cali-
brated with sucrose (AOAC, 1980). pH adjustments in
apple and orange juices prepared according to the
expected Brix (SS or % by weight of sucrose) values
were carried out by using 1N H2SO4 (Merck, Darms-
tadt, Germany) and 1N NaOH (Merck) while the same
adjustments in the MEB were carried out by using 1N
HCl (Merck) (Silva et al., 1999). After adjusting the SS
and pH values, 50 mL of apple juice, orange juice and
MEB solutions were sterilised in 100 mL glass tubes.
Thermal processing study
The thermostat controlled water bath (GFL Model
02.275, Germany) used for thermal processing was
adjusted to the intended temperature (85, 90, 95 and
100 C). Once it reached the intended temperature, the
bottles containing 50 mL apple juice, orange juice or
MEB solutions were placed in the bath. Upon waiting
nearly 25–30 min for the bottles to reach the intended
temperatures, 1 mL spore suspension was added and
mixed with vortex. Then the thermal processing was
applied to this samples (Acar & Cemeroglu, 1998; Vieira
et al., 2002).
Microbiological analysis
The number of spores in 1 mL of the media was
determined at the each thermal processing study.
For determining spore numbers of media, 1 mL of
samples were taken from sample by using sterile-end
automatic pipettes at the beginning and each
specific thermal treatment periods. Each taken sample
was added into the tubes containing 9 mL sterile
physiological water and then heat-treated samples
were cooled down by dipping in the cold water. Later,
their dilutions were prepared in appropriate levels
(Acar & Cemeroglu, 1998; Vieira et al., 2002). Approxi-
mately 50 lL samples of appropriate dilutions were
spread on BAT agar in duplicate and incubated at 45 C
for 24 h. The colonies in the petri plates were counted,
and the number of spores (cfu mL)1) in suspensions
after each heating period was calculated. Before heat
100
treatment, colony counts were recorded as the initial
25
number (No). Numbers of surviving bacteria after
2
heat treatment were recorded as survival bacteria (N)
(Sahbaz et al., 1996).
 2009 The Authors. Journal compilation  2009 Institute of Food Science and Technology
 Thermal resistance of A. acidoterrestris spores G. Ceviz et al. 1773

D and z-values
D-value is the thermal processing duration needed for
inhibiting 90% of micro-organisms in the media under
constant temperature. On the other hand, z-value is
defined as the temperature value which needs to be
increased to provide a decimal reduction (90%) in the
D-value (Sahbaz et al., 1996). In order to determine these
values, first of all, the number of bacteria (N) surviving in
the fixed temperatures of each sample and as a result of
the thermal processing carried out on regular intervals
was determined. The logarithms of the number of
surviving bacteria were graphed against the duration of
thermal processing and thus ‘survivor curves’ were
obtained. D-values were calculated from the slope of
the regression line obtained from the linear portion of the
survival curve. z-values were determined from the
regression lines obtained by plotting log D-values against
the corresponding temperatures (85, 90, 95 and 100 C)
(Unluturk & Turantas, 2003; Ozkan & Cemeroglu, 2005).

Results and discussion

The effects of thermal processing applications on the
inhibition of A. acidoterrestris spores were determined
under different temperature and duration conditions in
apple juice, orange juice and MEB. The survival curves
obtained for samples with SS = 10Brix and 3.5 pH are
showed in Fig. 1a, b and c. The survival curves for other
pH and SS values were obtained with similar methods
(data not shown). D-values calculated from the slopes of
survival curves and z-values found in accordance with
D-values are showed together in Table 2. Medium
temperature and the duration of the thermal processing
can be easily noticed to have significant effects on the
inhibition of A. acidoterrestris spores when Fig. 1a, b
and c are examined. It has been determined that the
temperature increase is very effective on the speed of the
inhibition and the inhibition occurred more speedily
under especially 100 C media.
It has been found that the effect of media on D-values Figure 1 Effect of temperature and time on the reduction of Alicy-
clobacillus acidoterrestris strain DSM2498 spores in apple juice (a),
obtained from apple juice, orange juice and MEB media
orange juice (b) and malt extract broth (c) at [soluble solids
is not significant (P > 0.05). On the other hand, if (SS)] = 10Brix, pH 3.5 and 85 C (¤), 90 C ( ), 95 C ( ), 100 C
D-values in Table 2 obtained for each media are (·).
specifically examined separately for each SS, it can be
easily concluded that the SS in the media does not affect value, however, the change in SS value do not have an
D-values in a specific way or direction. It has been obvious effect on D-value at 97 C. According to the
observed that D-value decreases as long as SS value literature, the water activation degree of the media is an
increases in apple juice and orange juice media whilst effective factor of the thermal resistance of micro-
D-values increases when SS value increases in MEB organisms and this resistance increases in parallel with
media. However, as a result of the statistical analyses, the decrease in the water activation value in the media
these changes in D-values depending on the SS value (Unluturk & Turantas, 2003). Besides, according to the
have been found to be of insignificant levels (P > 0.05). results of the study carried out by Splittstoesser et al.
Silva et al. (1999) reported that D-values of A. acido- (1998), the increase in the SS value up to a certain limit
terrestris spores in MEB media increase under low do not affect the thermal resistance of A. acidoterrestris
temperature conditions depending on the increase of SS spores, but it has been reported that high levels of SS

 2009 The Authors. Journal compilation  2009 Institute of Food Science and Technology International Journal of Food Science and Technology 2009
1774 Thermal resistance of A. acidoterrestris spores G. Ceviz et al.

Table 2 D and z-values of Alicyclobacillus acidoterrestris strain DSM2498 spores heated in apple juices, orange juices and MEB of different pH and
SS values

D-values (min)*

Heating medium SS pH 85 °C 90 °C 95 °C 100 °C z-values (oC)*

Apple juice 10 3.5 117.6 ± 9.48a 32.8 ± 1.84b 20.8 ± 1.98b 4.1 ± 0.64c 10.9 ± 0.58b
4.0 105.3 ± 5.52a 36.1 ± 4.53b 27.8 ± 1.70b 11.1 ± 2.55c 16.4 ± 0.81a
20 3.5 103.1 ± 6.43a 52.9 ± 3.96b 41.7 ± 6.51bc 6.2 ± 1.13d 13.3 ± 1.25a
4.0 94.3 ± 4.45a 43.7 ± 2.97b 18.5 ± 2.83c 3.8 ± 0.78d 10.9 ± 0.88a
Orange juice 10 3.5 93.5 ± 5.37a 36.5 ± 2.47b 20.8 ± 3.11c 3.3 ± 0.57d 10.9 ± 0.44a
4.0 120.5 ± 11.46a 30.9 ± 3.61b 20.8 ± 1.27bcd 2.3 ± 0.14d 9.4 ± 0.32a
20 3.5 88.5 ± 8.06a 35.3 ± 3.11b 16.7 ± 2.97c 4.0 ± 0.35dc 11.4 ± 0.30a
4.0 123.5 ± 5.09a 36.1 ± 2.69b 16.7 ± 1.98c 2.7 ± 0.28d 9.4 ± 0.18a
MEB 10 3.5 82.6 ± 5.52a 20.7 ± 2.97b 10.4 ± 2.12bcd 5.2 ± 0.64d 12.8 ± 0.74a
4.0 104.2 ± 3.82a 24.9 ± 1.98b 11.1 ± 2.26c 2.7 ± 0.42d 9.8 ± 0.37a
20 3.5 73.5 ± 4.31a 33.0 ± 2.76b 13.9 ± 0.57c 3.1 ± 0.42d 11.1 ± 0.39a
4.0 107.5 ± 9.48a 30.4 ± 3.18b 23.8 ± 1.70b 2.9 ± 0.21d 10.7 ± 0.13a

MEB, malt extract broth; SS, soluble solids.

*D-values represented (in the same columns and lines) by the same letters in 3.5–4.0 pH and different temperatures for each in the table above are not
statistically different from each other (P > 0.05). Similarly, z-values represented with the same letters for two different pH values in each SS are not
statistically different from each other (P > 0.05).

values contribute to the increase of thermal resistances.
As we exceed SS = 20Brix in our study, we do not
observe any difference stemming from SS change in
thermal resistance; and this fact do not contradict to the
literature.
One of the factors having an effect on the thermal
resistances of micro-organisms is the pH value of the
media. Although there is a general acceptance in
literature that the thermal resistance of micro-organisms
decreases as long as they become distanced from neutral
pH (Unluturk & Turantas, 2003), no statistical differ-
ence has been found between D-values obtained for 3.5
and 4.0 pH in apple juice and orange juice media
(P > 0.05). Nevertheless, it has been reported that
D-values obtained for 3.5 and 4 pH and SS = 20Brix
under 95 C in MEB media are statistically significantly
different (P < 0.05) while D-values for 3.5 and 4 pH
under different temperatures are not statistically differ-
ent from each other (P > 0.05). The results of the study
conducted by Murakami et al. (1998) on the effect of
thermal processing temperature and pH on the thermal
resistance of A. acidoterrestris AB-1 spores support the
results of our study. These researchers determined that
the effect of pH, in general terms, on the thermal
resistance of A. acidoterrestris AB-1 spores is not
statistically significant (P > 0.05); however they found
that D-values obtained for 5.0 pH and 8.0 pH under
only 92 C are significantly different from each other
(P > 0.05) as a result of the study they conducted in
Mcllvaine buffer, in 3–8 pH interval for six different pH
values and different temperatures (88, 90, 92 and 95 C).
Pontius et al. (1998) reported that pH at 91 C is
effective on the D-values of A. acidoterrestris spores in
model and fruit juice media, however pH is not effective
at 97 C. Similar results were achieved in the study
carried out by Silva et al. (1999). Researchers put
forward that the increase in pH under model 85 C
increase D-value of A. acidoterrestris (strain, NCIMB
13137) spores, too, in MEB media while D-value is not
affected by the increase of pH at 97 C. Similarly, Palop
et al. (2000) explained that there is no specific effect of
pH on the thermal resistance of Alicyclobacillus acido-
caldarius in Mcllvaine citrate–phosphate buffer with pH
value of 4.0 and 7.0.
Temperature has been found to be the most important
factor having effects on D-value, which is the main
indicator of the thermal resistance of A. acidoterrestris
spores among SS, pH and temperature values. D-values
have been observed to decrease specifically depending on
the increase of temperature and these values have been
found significantly different from each other (P < 0.05).
The decrease in D-value depending on the increase in the
temperature is shown in Fig. 2a–c more clearly. Whereas
D-values show a remarkable decrease in parallel with the
increase in the temperature, this decrease occurs in greater
rates in 85 C–90 C temperature interval. The decrease
in D-values has determined to decrease proportionally in
90–100 C interval to 85–90 C interval. In the study
carried out by Nakauma et al. (2004) on the thermal
resistance of A. acidoterrestris spores, the decrease in the
number of micro-organisms depending on the tempera-
ture was examined in two phases. D and z-values were
calculated separately for each phase depending on these
two phases. It was reported in the first logarithmic phase
that there was a rapid decrease in the number of micro-
organisms while in the second phase the resistance of

International Journal of Food Science and Technology 2009  2009 The Authors. Journal compilation  2009 Institute of Food Science and Technology

Figure 2 Decimal reduction time curves for Alicyclobacillus acidoter-
restris strain DSM2498 spores in apple juice (a), orange juice (b) and
malt extract broth medium (c) at [soluble solids (SS)] = 10Brix and
pH 3.5 (d), SS = 10Brix and pH 4.0 ( ), SS = 20Brix and pH 3.5
( ), SS = 20Brix and pH 4.0 (·).
micro-organisms was reported to increase and thus
raising the levels of D and z-values.
It is stated in literature that D-values of A. acidoter-
restris spores and other spores belonging to Alicycloba-
cillus species decrease in parallel with the increase in the
temperature of thermal processing; however, D-values
 2009 The Authors. Journal compilation  2009 Institute of Food Science and Technology
Thermal resistance of A. acidoterrestris spores G. Ceviz et al. 1775
determined for similar temperatures are much more
different than each other. Splittstoesser et al. (1994)
obtained the following results at the end of the study
conducted with A. acidoterrestris VF spores in apple
juice media: D85 = 56 min, D90 = 23 min,
D95 = 2.8 min and z = 7.7 C. In addition, D95 value
was reported to be between 2.1 and 2.5 in the apple juice
media of Alicyclobacillus isolated in Australia (Jensen,
2000). These findings are similar to the results of
Splittstoesser et al. (1994). On the other hand, in a
similar study related to the same issue, it was determined
that no significant decreases occurs in the number of
A. acidoterrestris spores in the apple juice media by
applying 1 min of temperature of 90 C (Lee et al.,
2002). In addition, Palop et al. (2000) found D-value of
A. acidocaldarius in distilled water media as D110 = 3.7,
D115 = 0.48, D120 = 0.11, D125 = 0.024 min. Another
study conducted in relation to this issue puts forward
that A. acidoterrestris spores are highly resistant to heat
in the orange juice media with 3.5 pH and SS = 9Brix:
D85 = 60.8–94.5 min, D90 = 10.0–20.6 min,
D95 = 2.5–8.7 min depending on the strain (Eiroa
et al., 1999).
As understood from the above statements, the values
determined by the researchers are different from both
other studies and from the results of this study. This
difference can be the result of the species and strain
difference of examined samples and their differently
adjusted thermal processing temperatures.
It has been also determined that the effect of media on
z-values obtained for apple juice, orange juice and MEB,
like in D-values, is not significant (P > 0.05). When the
evaluation has been made by taking into account each
thermal processing media (apple juice, orange juice and
MEB) separately, the effect of the change in SS value on
the z-value is found to be insignificant in MEB
(P > 0.05). Although the SS change has been reported
to be significantly effective (P < 0.05) on z-value in only
orange juice media with pH of 4.0, z-value has been
found to be lower in SS = 20Brix than in
SS = 10Brix and this fact contradicts to the general
expectation. These results obtained as the consequence
of our study are in parallel with the findings of
Splittstoesser et al. (1998). In the study, in which
researchers aimed to determine the thermal resistance
of A. acidoterrestris WAC spores under 85, 90 and 95 C
temperatures in Concord-type grape juice with 16, 30
and SS = 65Brix, they found a decrease in z-value in
SS = 30Brix (z = 6.6 C) when compared with
SS = 16Brix (z = 6.9 C) whereas they determined a
higher z-value in SS = 65Brix (z = 7.4 C) when
compared with both SS = 16Brix and SS = 30Brix.
These results obtained by the researchers support the
findings of our study as they suggest that the expected
result ‘the more SS the higher thermal resistance’ is
achieved in considerably high SS values.
International Journal of Food Science and Technology 2009
1776 Thermal resistance of A. acidoterrestris spores G. Ceviz et al.
When the effect of pH on z-value is investigated again
by taking into account each sample separately, it is
determined that z-value shows an increase depending on
the increase in pH value in SS = 10Brix apple juice and
this change is statistically significant (P < 0.05). Other
z-values obtained for 3.5 and 4.0 pH in all of the thermal
processing media are not significantly different from each
other (P > 0.05) (Table 2). Pontius et al. (1998) stated
that z-values do not show specific changes depending on
pH. A similar result was reported by Murakami et al.
(1998). In order to determine the effect of pH on the
thermal resistance of A. acidoterrestris AB-1 spores the
researchers carried out a study in which they experi-
mented six different pH values in pH 3–8 interval in
Mcllvaine buffer and as a result reported that z-values
changed little between 7.1 C and 6.4 C and they even
found that the z-values found in this interval were the
same values. These results put forward by various
researchers are in parallel with the results of our study.
As a result of the studies carried out with the aim of
determining the thermal resistances of Alicyclobacillus, it
has been observed that D and z-values obtained by
many researchers for different Alicyclobacillus species
and strains, different SS, pH and temperature conditions
show great variations (McIntyre et al., 1995; Pontius
et al., 1998; Eiroa et al., 1999; Palop et al., 2000; Silva &
Gibbs, 2001, 2004; Vieira et al., 2002). However,
according to both literature results and the results of
this study, the mentioned factors having effects on the
thermal resistance are not always significant and in the
expected direction.
Conclusions
According to the results obtained in this study, spores
can be determined to be highly resistant to the temper-
ature in thermal processing applications for the media
with broader intervals in terms of SS and pH values.
A. acidoterrestris, though non-pathogenic, generally
encountered as a spoiling factor and can survive during
pasteurisation in fruit juices and concentrated fruit
juices. A. acidoterrestris should be regarded as the
target micro-organism in the pasteurisation process.
Because of the pasteurisation norms cannot guarantee a
complete inactivation of A. acidoterrestris spores, the
final product should be kept at temperatures below
25 C to prevent their growth, and thus any risk of
spoilage.
Acknowledgments
The authors would like to thank to Dr K. Savas Bahceci
(Hacettepe University, Ankara, Turkey) for supplying
the culture. Additionally, authors would like to thank
Pamukkale University Scientific Researches Unit (Pro-
ject no. 2005FBE003) for financial support.
International Journal of Food Science and Technology 2009
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