Food Research International 101 (2017) 17–23

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Food Research International

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Microbial inactivation and evaluation of furan formation in high hydrostatic MARK

pressure (HHP) treated vegetable-based infant food
Gulcin Kultura, N.N. Misrab, Francisco J. Barbac,⁎, Mohamed Koubaad, Vural Gökmene,
Hami Alpasf
a
Republic of Turkey Ministry of Food, Agriculture and Livestock, Eskisehir Road 9th Km Lodumlu, Ankara, Turkey
b
GTECH, Research & Development, General Mills, Mumbai, India
c
Universitat de València, Faculty of Pharmacy, Preventive Medicine and Public Health, Food Science, Toxicology and Forensic Medicine Department, Nutrition and Food
Science Area, Avda. Vicent Andrés Estellés, s/n 46100 Burjassot, València, Spain
d
Sorbonne Universités, Université de Technologie de Compiègne, Laboratoire Transformations Intégrées de la Matière Renouvelable (UTC/ESCOM, EA 4297 TIMR),
Centre de Recherche de Royallieu, CS 60319, 60203 Compiègne Cedex, France
e
Food Quality and Safety (FoQuS) Research Group, Department of Food Engineering, Hacettepe University, 06800 Beytepe, Ankara, Turkey
f
Department of Food Engineering, Middle East Technical University, Ankara, 06800, Turkey

A R T I C L E I N F O A B S T R A C T

Keywords: The inactivation of pathogenic and spoilage bacteria as well as the formation of food processing contaminants
High pressure processing (e.g. acrylamide, furan, etc.) in infant foods is of utmost importance for industry, consumers as well as regulatory
Vegetable puree bodies. In this study, the potential of high hydrostatic pressure (HHP) for microorganism inactivation including
Mesophilic bacteria total mesophilic aerobic bacteria (TMA) and total yeasts and molds (TYM) at equivalent processing conditions,
Yeast
as well as its effects on furan formation in vegetable-based infant food was evaluated. The process parameters
Mold
evaluated were combinations of pressures (200, 300, and 400 MPa), temperatures (25, 35, and 45 °C), and
Furan
Food safety treatment times (5, 10, and 15 min). Pressure, time and temperature had a significant influence on both TMA
and TYM inactivation of vegetable-based infant foods, observing a significant reduction in both microbial po-
pulations when all the factors were increased, although the extent of reduction was clearly influenced by the
type of microorganism. A synergism between pressure, time and temperature was observed for the reduction of
both TMA and TYM populations and it was found that HHP at 400 MPa resulted in a complete inactivation of
TMA as well as TYM after 15 min of treatment at 45 °C. The furan content in all HHP treated samples was found
to be below the limit of detection. Thus, HHP treatment could be considered as a potential alternative to thermal
processing of vegetable-based infant foods.

1. Introduction National Institutes of Health, furan is carcinogenic to rats and mice,

Furan is a heterocyclic organic compound, consisting of a five-
membered aromatic ring with four carbon atoms and one oxygen. The
occurrence of furan in foods, the mechanisms leading to furan forma-
tion and analytical methods for detection have been reviewed in the
past (Crews & Castle, 2007; Wenzl, Lachenmeier, & Gökmen, 2007; Van
Lancker, Adams, Owczarek-Fendor, De Meulenaer, & De Kimpe, 2010).
This compound is formed during thermal treatment of food products.
Furan has been shown to be carcinogenic in animal laboratory studies
by Gill et al. (2011) and the International Agency for Research on
Cancer (IARC), thus constituting a chemical hazard (IARC, 2012). Per
the toxicology and carcinogenesis studies of furan made by U.S.
showing a dose-dependent increase in hepatocellular adenomas and
carcinomas (NTP, 1993). Furan induced carcinogenesis is suspected to
be either genotoxic mediated (Burka, Washburn, & Irwin, 1991; Byrns,
Predecke, & Peterson, 2002) or metabolite induced cell proliferation
and uncoupling of mitochondrial oxidative phosphorylation
(Kedderis & Ploch, 1999).
In the past, the US Food and Drug Administration (US FDA) had
released a report on the occurrence of furan in many foods that are
subjected to thermal processing, especially canned and jarred foods (US
FDA, 2009). Coffee and ready-to-eat baby foods are reported to contain
the high concentrations of furan among most ready-to-eat foods (EFSA,
2011). Historical FDA data on furan reveals that furan contents in

⁎
Corresponding author at: Nutrition and Food Science Area, Preventive Medicine and Public Health, Food Sciences, Toxicology and Forensic Medicine Department, Faculty of
Pharmacy, Universitat de València, Avda. Vicent Andrés Estellés, s/n, 46100, Burjassot, València, Spain.
E-mail address: francisco.barba@uv.es (F.J. Barba).

http://dx.doi.org/10.1016/j.foodres.2017.07.064
Received 6 February 2017; Received in revised form 25 July 2017; Accepted 26 July 2017
Available online 27 July 2017
0963-9969/ © 2017 Elsevier Ltd. All rights reserved.
G. Kultur et al.
commercially available infant foods typically range between 3.9 and
26.9 ppb (US FDA, 2009). It is interesting that the menace of furan
formation is limited mainly to commercially sterilized baby foods,
while freshly cooked home-made baby food is generally furan-free (Van
Lancker et al., 2010). In light of this, the use of mild processing tech-
nologies, which can avoid the use of high temperatures, can be a useful
tool to control furan level in baby foods (Barba, Terefe, Buckow,
Knorr, & Orlien, 2015; Sevenich et al., 2013).
Among mild processing technologies, high hydrostatic pressure
(HHP) processing has emerged as the most relevant one for food pre-
servation mainly due to its ability to inactivate microorganisms (> 5-
log reduction) (Baptista, Rocha, Cunha, Saraiva, & Almeida, 2016;
Georget et al., 2015; Moreirinha, Almeida, Saraiva, & Delgadillo, 2016;
Rendueles et al., 2011) and effectively control certain enzymes without
destroying the nutritional and sensory components that are normally
affected during heat treatment (Castro, Saraiva,
Domingues, & Delgadillo, 2011; Misra, Kadam, & Pankaj, 2011; Terefe,
Buckow, & Versteeg, 2014). A remarkable point for HHP processing is
the use of 3-D thinking, meaning that it is possible to control three
processing parameters (pressure, temperature and time), which offer a
high versatility in the process design (Barba, Esteve, & Frígola, 2012;
Barba, Parniakov et al., 2015; Barba, Terefe et al., 2015).
At this stage of development, there is a need to optimize HHP
processing conditions to achieve a balance between safety, food quality,
and health. Knowing the influence of HHP factors on microbial in-
activation is of great importance for both food researchers and food
industry as these are the key to develop innovative and effective pro-
cesses and products. However, it is also necessary to evaluate the in-
fluence of process variables on the formation of processing con-
taminants (e.g. furan), which can have an important effect on infants'
health and determine the consumer's acceptance.
To the best of our knowledge, there is a lack of information in the
available literature on the impact of HHP on microorganism inactiva-
tion/reduction on vegetable-based infant foods. Although, some pre-
vious studies have evaluated the formation/mitigation of furan and
MCPD-esters in different food systems after applying HHP (Sevenich
et al., 2013, 2014), the authors only evaluated the effect of a single
pressure value (600 MPa) combined with very high temperatures (90,
105, 110, 115, and 121 °C). No attempts have been made to system-
atically evaluate the influence of HHP processing conditions (pressure,
time and temperature) on microbial inactivation and furan formation/
mitigation in vegetable-based infant foods. Therefore, in the present
work, we focus on background microflora inactivation and furan for-
mation in infant foods, because infants and toddlers are the most sus-
ceptible consumer groups (Suk, Murray, & Avakian, 2003) and because
relatively high amounts of furan were previously detected in baby foods
(Lachenmeier, Reusch, & Kuballa, 2009; US FDA, 2009).
In the present work, a systematic study was conducted to evaluate
the effects of pressure, time and temperature on (i) total mesophilic
aerobic (TMA) bacteria, (ii) total yeasts and molds (TYM), and (iii)
furan formation, in vegetable-based infant foods. The results of our
multi-parameter study were analysed using multiple linear regression to
assess the relative contribution of each process variable.
2. Materials and methods
2.1. Samples and HHP treatments
The vegetable-based baby food was based on a mixture of water,
carrot, white cabbage, potato, marrow (a sweet variety of zucchini),
rice flour, celery root, sugar, tomato juice concentrate, salt and sun-
flower oil. The samples for experimentation were kindly supplied by
baby foods division of Hero Group, Turkey (Herobaby, 2017). The in-
itial mesophilic aerophilic bacteria population was 6.8 log10/g, while
that for total yeasts and molds was 5.8 log10/g. All samples for ex-
periment were drawn from the same pool of product, thereby ensuring
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Food Research International 101 (2017) 17–23
uniform initial microbial loads. The samples were deaerated by va-
cuuming. For pressure treatment, samples were carefully filled into
20 mL plastic bottles (LP Italiana SPA, Italy) to avoid any air bubbles,
undesirable in HHP process.
HHP treatment was performed in a 760.0118 type pressure equip-
ment supplied by SITEC-Sieber Engineering AG, Zurich, Switzerland.
The vessel had a volume of 100 mL with an inner diameter of 24 mm
and length of 153 mm. A built-in heating-cooling system (Huber
Circulation Thermostat, Offenburg, Germany) was used to maintain and
control the required temperature. The temperature in the vessel was
monitored using a type K thermocouple. The vessel was filled with a
pressure transmitting medium consisting of distilled water.
The increase in temperature originating from adiabatic heating was
calculated to be between 4 and 5 °C. The pressurization was applied to
the samples at a pressure of 200, 300, 400 MPa, temperature of 25, 35,
45 °C, for 5, 10, 15 min. Come up and pressure release times were not
considered for the HHP application times reported in the study. HHP
conditions were decided with respect to the literature research (Barba,
Koubaa, do Prado-Silva, Orlien, & de Souza Sant'Ana, 2017; Georget
et al., 2015). Pressurization rates were 400 MPa/min for 200 MPa,
360 MPa/min for 300 MPa and 340 MPa/min for 400 MPa (Günlü,
Sipahioǧlu, & Alpas, 2014; Subasi & Alpas, 2017). Pressure-treated
samples were stored at − 18 °C until performing the chemical and mi-
crobiological analysis. It may be noted that when time is not a con-
straint, performing microbiological analysis on the same day is re-
commended to avoid any sub-lethal injury to cells (Banwart, 1989; Jay,
Loessner, & Golden, 2005); however, in the present study the relative
recovery of TYM/TMA would be closely comparable for heat treated
and HHP processing, thus sufficiently serving the objective of com-
paring the two processing approaches. Samples to be treated thermally
were filled to the jars at 80 °C and subsequently pasteurized at 105 °C
for 10 min. The setup of the experimental design is represented in
Fig. 1.
2.2. Microbial enumeration
In this work, reductions in background microflora, i.e. total meso-
philic aerophiles and yeasts/molds, resulting from HHP processing
were studied, as these organisms are primary contributors to spoilage of
vegetable based infant foods under most practical conditions. The mi-
crobiological enumerations were carried out as follows.
2.2.1. Total mesophilic aerobic (TMA) bacteria
One gram of pressure-treated sample was suspended in 0.1% pep-
tone water. Inoculations were performed from 1:10 dilution. 1 mL
suspension of the baby food samples were surface plated on pre-poured
Plate Count Agar (Merck, Darmstadt, Germany) in three plates (0.3, 0.3
and 0.4 mL). After the incubation at 37 °C for 48 h, colony enumeration
was achieved.
2.2.2. Total yeasts and molds (TYM)
The procedure was similar to the determination of total mesophilic
aerobic bacteria except the medium and the incubation time. 1 g of
pressure-treated sample was suspended in 0.1% peptone water.
Inoculations were performed from 1:10 dilution. 1 mL suspension of the
baby food samples were surface plated on pre-poured Potato Dextrose
Agar (Merck, Darmstadt, Germany) in three plates (0.3, 0.3 and
0.4 mL). 14 mL of 10% tartaric acid was put to 1 L medium to avoid the
bacterial growth (pH 3.5). After incubation at 25 °C for 120 h, colony
enumeration was achieved.
2.3. Determination of furan content
Untreated, thermally- and HHP-treated vegetable-based baby food
were transferred to vials (Supelco, Bellefonte, PA, USA) and kept at 4 °C
to avoid any loss of furan due to its high volatility. Aluminium crimp
G. Kultur et al.
seals with Teflon-faced silicon septa were used in analyses (Agilent,
Waldbronn, Germany). About 5 g of sample was put into 20 mL head-
space vial and a Carboxen-PDMS SPME fiber (Supelco, Bellefonte, PA,
USA) was introduced. The sample with Carboxen-PDMS SPME fiber was
equilibrated to 40 °C in an incubator for 30 min. After equilibration,
SPME fiber was introduced into the injection port of GC and kept for
5 min for desorption. Chromatographic separation was achieved in a
capillary column (24 m × 320 μm, 20 μm) of HP-PLOT-Q connected to
Agilent 5973 N GC/MS (Agilent, USA). Helium gas at a rate of 2 mL/
min (60 cm/s) was used as a carrier gas. Column temperature program
was set as follows: 100 °C for 5 min, ramped to 200 °C at a rate of 10 °C/
min, followed by holding at 200 °C for 15 min, resulting in a total
analysis time of 30 min. Mass spectrometry was executed in the range
of m/z 20–200 AMU scan. Quantifier/qualifier ions for furan and d4-
furan were set as 68/39 and 72/43 m/z. The elution time of furan was
found to be 11 min. For calibration curve, external standard used was
furan (minimum purity 99%, Fluka Chemie GmbH, Switzerland), and
while 50 ng/g d4-furan (minimum purity 99%, Aldrich, USA) served as
internal standard. For quantitation, a calibration curve of furan was
prepared in the infant food covering the concentration range of
0–1000 ng/g. The limit of quantification (LOQ) for the method was
found to be 1 ng/g.
2.4. Statistical analysis
Two independent treatments were carried out in this study and the
mean results reported. A three-way analysis of variance (ANOVA) was
applied to the results obtained to verify whether there were significant
differences with respect to treatment pressure, temperature, and time,
and to ascertain possible interactions between the factors (differences at
p < 0.05 were considered significant). In order to capture the effects of
the three parameters, viz. pressure (P), treatment time (t), and tem-
perature (T) and their interactions, the experimental data for in-
activation of total mesophiles as well as yeasts and molds were fitted to
a quadratic function of the form-
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Food Research International 101 (2017) 17–23
Fig. 1. Schematic representation of the experimental setup.
Y = β0 + β1 P + β2 t + β3 T + β4 Pt + β5 PT + β6 tT + β7 P 2 + β8 t 2 + β9 T 2
where, βi are the coefficients of the variables, with β0 being the inter-
cept. This allowed to capture the purely quadratic effects as well as
interaction terms. We employed a stepwise linear fitting routine, which
allowed to retain only those terms which were significant (p < 0.05),
while simplifying the models without losing the accuracy of predictions
(Draper & Smith, 1981). The method uses a forward and backward
stepwise regression to determine the final model. An analysis of the
residuals revealed the suitability of the model for analysis of the ex-
perimental data. Statistical fitting was carried out in MATLAB (The
Mathworks, MA).
3. Results and discussion
3.1. Microbial inactivation
The rate and extent of microorganism inactivation after HHP differ
according to the processing conditions (pressure, time and temperature)
as well as to the food matrix and the type of microorganism (Georget
et al., 2015; Rendueles et al., 2011). The HHP treatments at 400 MPa
and 45 °C temperature for 15 min resulted in inactivation of mesophilic
aerophile bacteria by 6.8 log10 CFU/g, while that of yeasts and molds
by 5.8 log10 CFU/g. In this study, the relative impact of HHP processing
conditions (200, 300 and 400 MPa/25, 35 and 45 °C/5, 10 and 15 min)
on microbial inactivation (total mesophilic aerobic (TMA) bacteria and
total yeasts and molds (TYM)) was analyzed using multiple linear re-
gression (MLR). Multiple linear regression is a useful tool when con-
sidering the effect of more than one independent variable as is the case
with this study. Over the last years, MLR has been applied to analysis of
high pressure effects on virus inactivation (Calik, Morrissey,
Reno, & An, 2002), enzyme inactivation during high pressure proces-
sing in presence of CO2 (Truong, Boff, Min, & Shellhammer, 2002),
volatile formation during pressure assisted thermal treatment of milk
(Vazquez-Landaverde, Qian, & Torres, 2007), effects of e-beam
G. Kultur et al. Food Research International 101 (2017) 17–23

Table 1
Values of the significant coefficients of the quadratic model fitted to the inactivation data.

Coefficients Total mesophilic aerobes (TMA) Total yeasts and molds (TYM)

Estimate Standard error Estimate Standard error

Intercept (β0) − 10.251 1.784 − 5.281 1.290

Pressure (β1) 0.007 0.003 0.001 0.002
Time (β2) 0.522 0.116 0.783 0.169
Temperature (β3) 0.264 0.084 0.016 0.025
Pressure ∗ time (β4) 0.0003 – − 0.0004 0.002
Pressure ∗ temperature (β5) 0.0002 – – –
Time ∗ temperature (β6) − 0.007 0.001 − 0.006 0.002
Pressure2 (β7) – – – –
Time2 (β8) − 0.01 0.004 − 0.032 0.006
Temperature2 (β9) − 0.002 0.001 – –
R2 0.79 0.78
RMSE 0.39 0.567

irradiation on ready-to-eat food (Guillén-Casla, Rosales-Conrado, León-

González, Pérez-Arribas, & Polo-Díez, 2011), analysing the relation be-
Pressure: 200 to 400 tween wheat gluten and ACE inhibitory activity of hydrolysates (Zhang
et al., 2015), and quality changes in juices processed by non-thermal
technologies (Zarate-Rodriguez, Ortega-Rivas, & Barbosa-Canovas,
2000). MLR allows to assess the independent contribution of process
variables. The overall models described in the next sections were sta-
tistically significant (p < 0.05) with acceptable values of coefficients of
Time: 6.3 to 15
regression (R2) of ≈0.78, considering the involvement of three dif-
ferent independent process variables with complex interactions; similar
values of R2 have been reported previously for inactivation of aerobic
mesophilic bacteria and yeasts/molds during high pressure application
with the three independent variables selected in the study (Peñas,
Temperature: 25.2 to 45 Gomez, Frías, & Vidal-Valverde, 2008).

3.1.1. Model fitting for total mesophilic aerobic (TMA) bacteria

-1.4 -1.2 -1 -0.8 -0.6 -0.4 -0.2 For the inactivation data of total mesophilic aerobic (TMA) bacteria,
Main Effect the fitting routine allowed to eliminate several terms from the complete
second order model with interaction terms; the simplified model is
Fig. 2. Fixed effects plot depicting the contribution of individual variables towards in-
given as:
activation of the total mesophilic aerobic bacteria. The horizontal lines represent the
confidence intervals for the predictions. The units for parameters are as follows: Pressure
TMA = 7 × 10−3P + 0.522t + 0.264T + 3 × 10−4Pt + 2 × 10−4PT
in MPa, Time in min, and Temperature in °C. The effect corresponds to log10 TMA count,
with minus indicating decrease. − 7 × 10−3tT − 0.01t 2 − 2 × 10−3T 2 − 10.251 (1)

The values of the significant coefficients with estimation errors are

also provided in Table 1. In addition, the contribution of each of the
three variables towards the overall mesophile reduction effect is de-
picted in Fig. 2. The treatment temperature was found to have the most
Pressure: 200 to 400 significant effect on the inactivation of TMA bacteria, followed by
treatment time and finally pressure. The positive signs of the coeffi-
cients associated with the independent parameters (β1, β2, β3) suggest
that an increase in all three process variables (P, t, T) would result in a
greater inactivation of the TMA. Likewise, the negative sign of the
Time: 7 to 15 squared and interaction terms indicated a downward semi-parabolic
profile of the predicted TMA inactivation against these variables, as was
expected. Furthermore, the significant quadratic effect of temperature
and the interaction effects found in the present model is in agreement
with those previously reported for high pressure studies (Chakraborty,
Temperature: 25 to 45 Rao, & Mishra, 2015; Peñas et al., 2008). It also signifies that there is a
certain temperature within in the range studied, beyond which it con-
tributes to TMA inactivation. This is also valid for treatment time, as
depicted in Fig. 2. Peñas et al. (2008) also reported a positive effect of
-2 -1 0
increasing temperature from 10 to 40 °C on inactivation of aerobic
Main Effect mesophilic bacteria during high pressure application to vegetable
sprouts.
Fig. 3. Fixed effects plot depicting the contribution of individual variables towards in-
activation of the total yeasts and molds. The horizontal lines represent the confidence
intervals for the predictions. The units for parameters are as follows: Pressure in MPa, 3.1.2. Model fitting for total yeasts and molds (TYM)
Time in min, and Temperature in °C. The effects correspond to equivalent log10 count, The simplified form of the second order model for inactivation of
with minus indicating decrease.
yeasts and molds was found to be:

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G. Kultur et al. Food Research International 101 (2017) 17–23

Fig. 4. The reduction of (a) total mesophilic aerobic (TMA)

(a) 7 25 oC 7 35 oC 7 45 oC bacteria, and (b) total yeasts and molds (TYM) in the vegetable
based infant food as a function of time, at three different pres-
200 (MPa) 200 (MPa) 200 (MPa) sures. Error bars represent standard deviation.
6 300 (MPa) 6 300 (MPa) 6 300 (MPa)
400 (MPa) 400 (MPa) 400 (MPa)
TMA Count log10 CFU/g

5 5 5

4 4 4

3 3 3

2 2 2

1 1 1

0 0 0
0 5 10 15 0 5 10 15 0 5 10 15
Time (min) Time (min) Time (min)

(b) 25 oC 35 oC 45 oC
6 6 6
200 (MPa) 200 (MPa) 200 (MPa)
300 (MPa) 300 (MPa) 300 (MPa)
400 (MPa) 400 (MPa) 400 (MPa)
5 5 5
TYM Count log10 CFU/g

4 4 4

3 3 3

2 2 2

1 1 1

0 0 0
0 5 10 15 0 5 10 15 0 5 10 15
Time (min) Time (min) Time (min)

TYM = 1 × 10−3P + 0.783t + 0.016T + 4 × 10−4Pt − 6 × 10−3tT
+ 6 × 10−3t 2 − 5.281
The contribution of each of the three variables towards the reduc-
tion of yeasts and molds is depicted in Fig. 3. The overall model was
statistically significant (p < 0.05). The treatment time was found to
have the most significant effect on the inactivation of TYM, followed by
temperature and pressure. There could be several reasons for the
treatment time being a governing parameter, including a variability in
bacterial populations for sensitivity to pressures, mixed bacterial po-
pulations in samples, and/or yield points of the bacterial adaptation to
the pressure stress making them more susceptible (Chen,
Hoover, & Kingsley, 2005). From Fig. 3 it can be concluded that keeping
all other parameters at minimum (i.e. interpretation of all coefficients is
ceteris paribus), an increase in treatment time from 7 min to 15 min
would result in an average decrease in the TYM population by at least 2
log10. Similarly, increasing in isolation, the pressure or temperature,
would contribute to a decrease in TYM by less than 1 log10. The
quadratic effects of time indicated that a minimum holding time (of
2 min) is necessary before inactivation begins (see Fig. 3). Furthermore,
time played a greater role in inactivation of TYM than TMA, con-
sidering that the corresponding coefficient value is greater in the latter
case. Peñas et al. (2008) also reported that both linear and quadratic
(2) effects of time, as well as interactions between time and temperature
were significant during inactivation of yeasts and molds in mung bean
sprouts through high pressure application (100–400 MPa; 5–15 min;
10–40 °C).
3.2. Comparison of microbial reduction for TMA, total yeasts and molds
(TYM)
In order to compare the effect of HHP at equivalent conditions on
the inactivation of TMA and TYM, a three-way analysis of variance
(ANOVA) was conducted. As can be seen in Figs. 2–4, all the studied
factors (pressure, time and temperature) had a significant influence
(level of probability, p < 0.05) on TMA and TYM of the vegetable-
based infant foods after HHP, observing a decrease in the number of
TMA and TYM when pressure, treatment time and temperature were
increased (Fig. 4), and obtaining the highest reduction at 400 MPa/
45 °C/15 min, independently of the studied microbial population.
As can be observed in Figs. 2–4, synergism between pressure, time
and temperature was observed for the reduction of both TMA and TYM
populations, thus showing the special importance of optimizing pres-
sure-time-temperature processing conditions to reduce the level of

21
G. Kultur et al.
microbial populations in vegetable-based infant foods and, preserving
at the same time, the nutritional and quality attributes of these food
products. In this line, other authors have established the possibility to
reduce the temperature needed to achieve the same microbial in-
activation by increasing the process pressure (Buckow & Heinz, 2008).
Similar to the results found in the present study, the synergism between
pressure and temperature on microbial inactivation was previously
reported (Buckow & Heinz, 2008).
It should also be noted that in all cases, the extent of microbial
reduction found for each pressure-time-temperature combination dif-
fered according to the microbial population studied (TMA or TYM). As
can be seen in Fig. 4, TMA were more resistant to HHP compared to
TYM, especially when the HHP treatments were conducted at the lowest
temperature (25 °C). As it was shown previously by other authors, a
higher resistance to pressure has been observed for mesophilic bacteria
compared to yeasts and molds (Georget et al., 2015; Rendueles et al.,
2011), although there is a lack of information comparing equivalent
HHP processing conditions on the same food matrices. Pressure at a
sufficiently high level, induces enzyme inactivation, denaturation of
membrane protein and cell membrane rupture caused by a phase
transition of the membrane and its fluidity, thus resulting in in-
activating the microorganisms (Georget et al., 2015).
3.3. Furan content
Furan content in thermally processed vegetable-based infant for-
mula was found to be 5.84 ± 2.36 ng/g. This value is in agreement
with those reported by Sijia, Enting, and Yuan (2014). For all HHP
processed baby food samples, the furan contents were found to be
below the limit of quantification (i.e. 1 ng/g). This leads us to the
conclusion that HHP at the temperatures employed in our study does
not result in furan formation, thereby rendering HHP a suitable method
for ensuring microbial safety of baby foods without introducing pro-
cessing contaminants. Palmers et al. (2015) reported that high-pressure
high-temperature processing (HPHT) resulted in minimum furan levels
in the order of 1.5–2 ng/g in spinach purée. This was likely because the
treatments were carried out at temperatures higher than 100 °C, pro-
viding an environment conducive for chemical changes driving furan
formation. When used at ambient or sub-ambient temperatures, high
pressure application being a purely physical process, while deconta-
minating foods, is unlikely to result in furan formation. Palmers et al.
(2015) have also reported that a pressure level of 600 MPa did not af-
fect the rate of furan formation in spinach purée, as opposed to the
processing temperature and time. These results are also in agreement
with those of Sevenich et al. (2013) and Sevenich et al. (2014), who
found a significant reduction in furan formation in HHP-treated sar-
dines in olive oil and baby food purée, respectively, even under ster-
ilization conditions (600 MPa/121 °C).
4. Conclusions
The present work evaluated HHP application as an intervention for
inactivation of spoilage micro-organisms in vegetable based baby food
without significant furan formation. Pressures of 400 MPa at 45C, ap-
plied for 15 min was found to result in complete inactivation of total
aerobic mesophiles and yeasts/molds, with no furan detected. All the
factors, viz. pressure, temperature and time have significant roles in
inactivating micro-organisms. The rate of inactivation of total meso-
philes versus yeasts/molds differs depending on the HHP processing
conditions. In general, total mesophilic aerobic bacteria were observed
to be more resistant to HHP than yeasts and molds. For all the pro-
cessing conditions evaluated in the study, no furan was detected in the
treated samples. While, it was not an objective of this study to explore
the nutritional/quality profile of the HHP processed baby foods, it is
worthwhile mentioning that HHP allows to considerably retain nutri-
tional and quality parameters of fruit and vegetable products (Oey,
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Food Research International 101 (2017) 17–23
Lille, Van Loey, & Hendrickx, 2008; Wang et al., 2013). HHP is also
known to inactivate enzymes responsible for spoilage of fruit and ve-
getable products (Chakraborty, Kaushik, Rao, & Mishra, 2014). In con-
clusion, HHP can be a potential alternative to thermal processing for
inactivation of spoilage microbes in vegetable based infant foods, while
mitigating the challenge of furan, a food contaminant. Further studies
are required to be conducted at a pilot scale to prepare this technology
for adoption by the infant foods industry.
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