105
Universidad de Sonora
ISSN: 1665-1456
105
Journal of biological and health sciences
http://biotecnia.unison.mx
Volume XXV, Issue 2
*Author for correspondence: Juan Arturo Ragazzo Sanchez
e-mail: jragazzo@ittepic.edu.mx
Received: October 15, 2022
Accepted: January 03, 2023
Staphylococcus aureus inactivation and maintenance of macronutrients
of human milk by high hydrostatic pressure and spray-drying process
Inactivación de Staphylococcus aureus y mantenimiento del valor nutricional de la leche humana por
altas presiones hidrostáticas y secado por aspersión
Blanca Rosa Aguilar-Uscangaa, Montserrat Calderon-Santoyob, Maricarmen Iñiguez-Morenob,c, Josue Raymundo
Solis-Pachecoa, Angel Fonseca-Cantabrana,b and Juan Arturo Ragazzo-Sanchezb*
a Industrial Microbiology Laboratory. University Center of Exact Sciences and Engineering. University of Guadalajara. ,
Boulevard General Marcelino Garcia Barragan, Col. Olimpica, C.P. . Guadalajara, Jalisco, Mexico.
b Integral Laboratory for Food Research, National Technological Institute of Mexico / Technological Institute of Tepic, Tepic,
Nayarit, Mexico.
c Polytechnic University of the State of Nayarit, Carretera Tepic-Aguamilpa, Tepic, Nayarit, Mexico.
RESUMEN
Los bancos de leche humana (BLH) utilizan la pasteurización
y congelación, como principales métodos de conservación.
Sin embargo, su valor nutricional disminuye durante la des-
congelación y almacenamiento. El objetivo del presente es-
tudio fue evaluar el efecto de las altas presiones hidrostáticas
(HHP) y el secado por aspersión sobre los macronutrientes, la
calidad microbiológica e inactivación de Staphylococcus au-
reus en LH. Para ello, se realizó la cuanticación de proteínas,
lípidos, carbohidratos, cenizas, bacterias lácticas, mesólos
aerobios, coliformes, mohos, levaduras y S. aureus. Los re-
sultados mostraron que S. aureus y los grupos microbianos
evaluados fueron reducidos por debajo del límite permitido
por los BLH (<10 UFC/mL). Mientras que las concentraciones
de macronutrientes permanecieron sin cambio durante todo
el proceso de conservación. El uso de bra soluble durante el
proceso de secado permitió obtener un rendimiento mayor
al 99 %. El polvo mostró alta solubilidad y bajos niveles de
humedad y actividad en agua; las cuales son propiedades
deseables en los alimentos deshidratados. Por lo tanto, la
combinación de HHP y el proceso de secado por aspersión
demostró ser una alternativa para facilitar el manejo y mejo-
rar la calidad microbiológica de la leche humana.
Palabras clave: Leche humana, altas presiones hidrostáticas,
secado por aspersión, Staphylococcus aureus, métodos de
conservación
ABSTRACT
Human milk banks (HMB) use pasteurization and freezing
methods to preserve the milk, nevertheless, the nutritional
value of the human milk (HM) decreases during thawing
and storage. This study aimed to evaluate the eect of high
hydrostatic pressures (HHP) and spray-drying on macronu-
trients, microbiological quality, and inactivation of Staphylo-
coccus aureus on HM. The characterization of HM powder,
reduction of S. aureus, and modications in microbiological
viability, proteins, carbohydrates, lipids, and ashes were
assessed. The ndings demonstrated that while S. aureus
and all other examined microbial groups were undetectable,
macronutrient concentrations remained constant during the
entire conservation process. High yield (> 99 %) was achie-
ved thanks to the inclusion of soluble ber during the drying
process, and the produced powder displayed high solubility,
low moisture content, and activity water; which are desirable
properties in dried foods. Therefore, the combination of HHP
and the spray-drying process is an alternative to facilitate
handling, improve the microbial quality, allow the addition
of oligosaccharides, and maintain the nutritional value of HM
in HMB.
Keywords: Human milk, high hydrostatic pressure, spray
drying, Staphylococcus aureus, milk preservation
INTRODUCTION
Human milk (HM) is a biologically active food that contains
high-biological value compounds and immunological consti-
tuents, that play a vital role in the modulation of the immune
system in newborns and particularly in premature infants. For
this, human milk banks (HMB) promote, protect, and support
breastfeeding. Besides, these institutions are responsible
for collection, conservation, and distribution of milk to feed
premature infants and newborns with nutritional disorders
(Lowry et al., 1951; Slutzah et al., 2010; Solís-Pacheco et al.,
2019). The high availability of nutrients in fresh HM provides
an ideal culture media for several microbial groups, such as
Staphylococcus aureus, one of the most important bacteria
that can contaminate HM from origin, due to mastitis by
infection (Koenig et al., 2005; Mediano et al., 2017). Besides,
secondary contamination can occur due to unsatisfactory
hygienic and sanitary handling conditions during extraction
and storage (Mediano et al., 2017). To control HM microbial
populations, HMB implements several food conservation
techniques, with refrigeration and pasteurization as the most
widely studied methods (Koenig et al., 2005; Novak and Cor-
deiro, 2007; Slutzah et al., 2010; Wesolowska et al., 2019). In
line with this, HM can be stored at refrigeration temperature
DOI: 10.18633/biotecnia.v25i2.1872
106 Volume XXV, Issue 2
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106
(4 °C) for up to 96 h, without compromising its nutritional va-
lue and microbiological safety (Slutzah et al., 2010). Instead,
pasteurization is the most common technique for food con-
servation. Microbial inactivation by this process is based on
the maintenance of food temperature at 62.5 °C for 30 min
and rapid cooling at 5 °C. Pasteurization inactivates vegetati-
ve bacteria and most viruses, including human immunode-
ciency virus, herpes, and cytomegalovirus (Cavazos-Garduño
et al., 2016; Wesolowska et al., 2019). After pasteurization, HM
should be stored in airtight containers and frozen at -18 ºC for
preservation for four months. However, during this period,
the nutrients of the milk are not guaranteed (Wesolowska et
al., 2019).
Some innovative technologies have been proposed
to extend HM shelf-life preserving its nutritional value,
including high-temperature short-time pasteurization,
high hydrostatic pressure (HHP) processing, and microwave
irradiation (Wesolowska et al., 2019). However, once the HM
microbial load has been inactivated, HM handling is still
complicated due to the high volume required for storage.
Recently, the spray-drying process demonstrated to be an
alternative for reducing HM volume while maintaining more
than 95 % of the macronutrients (Solís-Pacheco et al., 2019).
During this process, polysaccharides, proteins, and ber can
be added as wall materials, preventing volatilization and
protecting encapsulated material against environmental
conditions (Afoakwah et al., 2012). In addition, the use of pre-
biotic bers in the HM drying process could act as a source
of oligosaccharides for newborns, who cannot obtain them
directly from the mother’s diet. The value of breastmilk oligo-
saccharides and dietary bers, in complementary nutrition
for the development of the infant’s microbiome with both
short- and long-term health complications, has been lately
highlighted (Çavdar et al., 2019). NUTRAFLORA® is a synthe-
tic short-chain fructooligosaccharide, made by enzymatic
reaction with sucrose. This prebiotic ber is low in calories
(1.5 kcal/g) and viscosity, and has stability at temperatures
used during the High-Temperature Short-Time process (>
169 ºC). Furthermore, is easy to dry and extrude, and does
not participate in Maillard reactions. Its high solubility makes
NUTRAFLORA® ideal for dry or liquid formulations (Ingredion,
2016). Hence, this study aimed to evaluate i) the eect of HHP
pretreatment and the spray-drying process on the preserva-
tion of the macronutrients of HM added with prebiotic ber,
ii) the inhibition of microorganisms naturally present in HM,
and iii) the reduction of S. aureus articially inoculated in HM.
MATERIALS AND METHODS
Biological material and ethical considerations
Hospital Civil of Guadalajara “Fray Antonio Alcalde” provide
frozen samples from its HMB. The samples were transported
in a cooler to the Laboratorio de Microbiología at Instituto
Tecnológico de Tepic for analyses. The Ethical Research
Committee approved this study on October 2019 No. HCG/
CEI-1225/17.
Inoculation of S. aureus in HM
S. aureus ATCC 25923 was provided by the Laboratorio de
Microbiología Industrial, Universidad de Guadalajara. The
stock cultures were kept at -80 °C in tryptic soy broth (TSB;
Becton Dickinson Bioxon, Le Pont de Claix, France) with 15
% (v/v) glycerol. Before experiments, 25 L of stock cultures
were transferred to 3 mL TSB and incubated at 35 °C for 18
h, then 25 L were added to new TSB and incubated at 35 °C
under static conditions for 18 h, to yield a nal concentration
of 106 CFU/mL of microorganisms at stationary phase. Bac-
terial cells were harvested by centrifugation (9390 × g for 5
min), washed twice in sterile saline solution [SS, 0.85 % (w/v),
sodium chloride at pH 7.0 ± 0.2], and centrifuged under the
same conditions. Then, the bacterial cells were resuspended
in the raw HM to a nal concentration of 106 CFU/mL (Bulut
and Karatzas, 2021).
HM conservation process
For the HHP process, HM was thawed and homogenized. Then
250 mL of non- and inoculated-samples with S. aureus were
vacuum-sealed in sterile FoodSaver® plastic bags (Newell
Brands, Hoboken, NJ, USA) and pressurized at 300 MPa (Avu-
re Autoclave Systems, Model LCIP402260NCEP1MLN, Eri, PA,
USA) under dierent conditions (Table 1). HM was dried using
a Mini Spray Dryer B-290 (Büchi, Flawil, Switzerland) with an
inlet temperature of 165 °C and an air outlet temperature
of 110 °C. The milk was fed with a 7 mm-diameter nozzle at
a constant ow rate (2 mL/min); during the whole process,
the sample was stirred at 100 rpm at 25 °C. Before drying,
the spray dryer was stabilized with sterile distilled water
(SDW) under the required operating conditions for 10 min.
Soluble ber (NUTRAFLORA® P-95 / L95-S, Ingredion, Ciudad
de México, Mexico) was used as wall material at 5 % (w/v).
The process yield was estimated (Eq. 1). Three replicates were
used for the test and the experiment was repeated twice.
(%) =
100
(1)
Where: WDS is the weight of dried milk and WSF is the
weight of solids in raw milk (Solís-Pacheco et al., 2019). The
powder was stored in PET/BOPP/PE zipper bags at 25 ± 2 °C.
Proximal analyses of HM and HM powder
Total protein was estimated by the method of Lowry et al.
(1951), lipid content was determined by the gravimetric
method described by Folch et al. (1957), and lactose con-
centration was obtained with Anthrone reagent following
Table 1. Conditions of high hydrostatic pressure treatments at 300 MPa.
Tabla 1. Condiciones utilizadas en los tratamientos de alta presión hidros-
tática a 300 MPa.
Treatment Temperature (°C) Time (min)
1 35 10
2 35 20
3 45 10
4 45 20
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the method of Dubois et al. (1956). Protein and lactose
quantication were carried out using calibration curves with
casein at 595 nm and lactose at 490 nm; respectively. On the
other hand, ashes were estimated by drying the samples in
an oven at 240 °C for 24 h and the content was expressed
as a percentage (Sögüt et al., 2013). Each determination was
carried out in triplicate and repeated once. The values were
compared with the data obtained from the tests carried out
using fresh HM. The methodological strategy realized in this
study is represented in Figure 1.
logarithmic microbial reduction was calculated (Eq. 2).
(2)
where N0 is the initial count and N is the survival count
after treatment (Coroller et al., 2006). Each test was carried
out in triplicate and repeated once.
Moisture content (MC) of spray-dried milk
The MC was gravimetrically determined (da Silva Carvalho et
al., 2016). For this, 2 g of sample were used and the thermo-
balance (Sartorius MA 35, Göttingen, Germany) was operated
at 105 °C, the test was nished until weights remained cons-
tant. The test was carried out in triplicate and repeated twice.
Water activity (aw)
The measurement was made by using 5 g of the sample in
the hygroscopic Aqualab 4TEV (METER Group, Inc., Pullman,
WA, USA). Calibration was carried out with a sodium chloride
solution at 0.75 aw. The test was carried out in triplicate and
repeated twice.
Powder milk solubility
The test was carried out according to the protocol proposed
by Cano-Chauca et al. (2005). Briey, 1 g of the powder was
added to 100 mL of distilled water and solubilized using a
Vortex mixer (Genie 2, Daigger, Wheaton, IL, USA) at 3200
rpm for 5 min. The sample was centrifuged (Hermle Z326K,
Wehingen, Germany) for 5 min at 3000 × g, then 25 mL of
the supernatant were transferred to pre-weighed Petri dishes
and immediately oven-dried at 105 °C for 5 h. The solubility
percentage was calculated by weight dierence. The test was
carried out in triplicate and repeated twice.
Statistical analyses
Factorial designs were carried out. The data were subjected
to analysis of variance, then; the post-hoc least signicant
dierence (LSD) Fisher test (p ≤ 0.05) was used for means
comparison. Analyses were performed by using Statgra-
phics Centurion XVI.I software (Statpoint Technologies, Inc.,
Warrenton, USA).
RESULTS AND DISCUSSION
Microbiological quality of HM
HM is an ideal medium for the growth of microorganisms due
to the high availability of nutrients. Some microorganisms,
such as S. aureus, E. coli, and Salmonella may contaminate
HM due to inadequate procedures during its extraction
and/or storage (Mediano et al., 2017; Novak et al., 2008). In
this study, the HM from the Bank of Hospital Civil de Gua-
dalajara showed low levels of aerobic mesophilic bacteria
(2.48 ± 0.25 Log10 CFU/mL), S. aureus (2.21 ± 0.17 Log10 CFU/
mL), and lactic acid bacteria (1.31 ± 0.44 Log10 CFU/mL).
Whereas coliforms, molds, and yeast were not detected by
the traditional counting plate. The initial counts of all tested
microorganisms are below the established limits for pasteu-
Figure 1. Methodological diagram of the eect of high hydrostatic pressure
and spray drying on human milk compositional and microbiological loads.
Figura 1. Diagrama metodológico del efecto de la alta presión hidrostática
y el secado por aspersión sobre la composición y carga microbiológica de la
leche humana.
Microbiological analyses in HM and milk powder
Quantication of mesophilic aerobic bacteria, total coliforms,
lactic acid bacteria, S. aureus, molds, and yeasts was carried
out in raw milk according to Mexican standards (NOM-111-
SSA1-1994, NOM-113-SSA1-1994, NOM-115-SSA1-1994,
NOM-184-SSA1-2002). Microbial counts were carried out on
Petri dishes with the corresponding culture media for each
microbial group and incubated under dierent conditions.
For yeast and molds, potato dextrose agar (PDA, Bioxon, Bec-
ton Dickinson and Company, Queretaro, Mexico) was used
and the plates were incubated for 3 and 5 days at 25 °C. The
culture media used for aerobic bacteria (tryptic soy agar, TSA;
Becton Dickinson, Le Pont de Claix, France), total coliforms
(Red violet bile agar, Merck, KGaA, Darmstadt, Germany),
lactic acid bacteria (MRS agar, Oxoid, Basingstoke, UK), and S.
aureus [Baird-Parker agar base (Difco, MI, USA) supplemented
with egg yolk and 100 pg/mL potassium tellurite] were incu-
bated at 37 °C for 24-48 h. Lactic acid bacteria were incubated
under anaerobic conditions. Besides, after each conservation
process yeast, molds, lactic acid bacteria, and coliforms
were estimated as previously described in non-inoculated
samples. Whereas, S. aureus was quantied in un- and treated
inoculated samples. For dried milk, rehydration was carried
out considering the moisture content of each sample. The
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108
rized milk or infant formula (‘NOM-131-SSA1-2012, Products
and services. Formulas for infants, continuation, and special
nutritional needs. Food and non-alcoholic beverages for
infants and young children. Provisions and specications’,
2012; ‘NOM-184-SSA1-2002, Products, and Services. Milk,
Milk Formula, and Milk Product Combined. Sanitary speci-
cations’, 2012; ‘NOM-243-SSA1-2010, Milk, milk formula,
combined milk product, and milk derivatives. Provisions and
sanitary specications. test methods’, 2010) considering that
the samples were still untreated. These results are in concor-
dance with previous reports of fresh milk obtained from the
same HMB (Cavazos-Garduño et al., 2016; Solís-Pacheco et
al., 2019; Aguilar-Uscanga et al., 2021). This fact allows us to
infer the establishment and compliance of good practices for
the obtention and handling of HM by the mentioned HMB.
This is of vital importance to maintain microbiological quality
and nutritional value, and to avoid infections in newborns via
the presence of microorganisms in the milk (Hartmann et al.,
2007). However, the microbial load in HM can be higher than
that obtained in this research due to some infections, such
as mastitis. In HM samples from women with mastitis, the
mean microbial load was 4.11 Log10 CFU/mL. The staphylo-
coccal group is the most frequently isolated (97.57 %), with
Staphylococcus epidermidis (91.56 %) and S. aureus (29.74 %)
as the most common species isolated from mastitis samples
(Mediano et al., 2017). Inadequate sanitation and handling
during milk extraction result in its contamination with E.
coli and Salmonella, while S. aureus contamination of HM
occurs in breastfeeding women as a result of clinical masti-
tis (Kaavya et al., 2021; Mediano et al., 2017). In addition, S.
aureus is present in the oropharynx and skin of humans, and
its presence in HM may be due to secondary contamination
or to unsatisfactory hygienic and sanitary conditions of the
extraction apparatus used (Mediano et al., 2017).
Proximal analysis of HM
The initial estimated values of lactose, lipids, proteins, and
ashes in raw HM were 7.36 ± 0.94, 2.46 ± 0.26, 1.65 ± 0.59
g/dL, and 0.26 ± 0.02 %; respectively. These results are in
concordance with previous reports for Mexican fresh HM
(Cavazos-Garduño et al., 2016; Solís-Pacheco et al., 2019)
and with the generally assumed HM composition (Boquien,
2018). After the HHP processing, the assessed components
were maintained at more than 95 % (Table 2). This behavior
agrees with the results of Solís-Pacheco et al. (2019) for spray-
dried HM under the same conditions. However, after the
spray-drying process lactose showed a signicant reduction
in comparison to raw milk (p ≤ 0.05), but without change in
comparison to unpressurized dry HM (p > 0.05). During the
spray-drying process, chemical and physical changes such as
crystallization and nonenzymatic browning as the Maillard
reactions can occur, reducing the lactose concentration
(Kinsella and Morr, 1984; Uscanga et al., 2021). Besides, the
concentration of macronutrients in HM can be inuenced by
various processes, such as storage, freezing, and thawing. It
has been reported that freezing, thawing, and spray-drying
processes can decrease the lipid content in HM by up to 9.0
%, attributable to its adherence to the wall of container and
equipment, incomplete homogenization, lipolysis, or lipid
peroxidation (Cavazos-Garduño et al., 2016; Chang et al.,
2012). In this research, the variation in lipid content after HHP
and spray-drying were not signicant (p ≤ 0.05; Table 3). The
increment in ashes content in the HM powder was attributed
to the use of soluble ber as wall material, which can contain
between 1.60 and 2.30 % of ashes depending on the source
and variety (Ragaee et al., 2012).
Microbial reduction and inactivation of S. aureus by con-
servation process
Microbial inactivation by HHP depends on several factors
including treatment conditions (surrounding media, tem-
perature, pressure, time, etc.) and microbial characteristics
(vegetive or spore cells, composition of the cell wall, physio-
logical state, and so on) (Kaavya et al., 2021; Salleh-Mack and
Roberts, 2007). The lethal eect of treatments on bacterial
cells increased as a temperature function (p ≤ 0.05; Table
4). This is in agreement with a previous report on the inac-
tivation of S. aureus and Bacillus cereus spores in HM after 4
cycles of 5 min at 38 °C and 350 MPa (Demazeau et al., 2018).
In line with this, it has been demonstrated that S. aureus
inactivation in cow’s whole milk at 20 °C occurs at >250 MPa
and 8 min under these conditions, a decimal reduction after
3.7 min at 300 MPa was reported (Erkmen and Karata, 1997).
The composition of the cell wall and shape of the bacteria
are related to their resistance. Spherical organisms ought
to be more resistant to crushing than those rod-shaped
Table 2. Eect of high hydrostatic pressure on the composition of human milk.
Tabla 2. Efecto de alta presión hidrostática en la composición de la leche humana.
Treatment Lactose (g/dL) Lipids (g/dL) Proteins (g/dL) Ashes (%) Humidity (%)
1 6.71 ± 0.69a2.89 ± 0.17a1.49 ± 0.10a0.27 ± 0.03a86.19 ± 0.50a
2 7.42 ± 0.06a2.44 ± 0.84a1.37 ± 0.64a0.20 ± 0.05a86.13 ± 0.07a
3 6.98 ± 0.63a1.90 ± 0.70a1.44 ± 0.22a0.31 ± 0.06a86.81 ± 0.68a
4 6.85 ± 0.60a2.34 ± 0.14a1.39 ± 0.43a0.30 ± 0.09a86.37 ± 0.34a
Values are expressed as mean ± standard deviation (n = 6). Parameters of raw human milk: lactose
7.36 ± 0.94 g/dL; lipids 2.46 ± 0.46 g/dL; proteins 1.65 ± 0.59 g/dL; and ashes 0.26 ± 0.02 %. Values
in the same column followed by dierent letters are signicantly dierent according to Fisher’s LSD
test at p ≤ 0.05.
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ones, being consistent with the idea that the lethal action
of pressure changes that accompany their passage through
liquids (Jacobs and Thornley, 1954). The HHP disrupts the
bacterial membrane, being Gram-positive more resistant
than Gram-negative bacteria (Costello et al., 2021; Huu et al.,
2021; Repine et al., 1981). This is related to the dierent pepti-
doglycan content, which is higher in Gram-positive (30-70 %)
than in Gram-negative (< 10 %) bacteria, making them more
resistant to pressure (Schumann, 2011). Microorganisms can
resist high pressures, due to membrane uidity. However,
the synergistic eect of the temperature contributes to the
destabilization of the outer and inner membrane, releasing
intracellular components (Huu et al., 2021; Kaavya et al., 2021;
Salleh-Mack and Roberts, 2007).
Spray drying is a technique widely used for milk powder
production. This process is not aimed to cause microbial inac-
tivation (Alvarenga et al., 2018). However, it showed a syner-
gistic eect in pressurized HM samples contributing to the
complete inactivation of S. aureus during the drying process,
because cells suered previous damage by the HHP (Bulut
and Karatzas, 2021) (Table 4). Temperature can have a cumu-
lative or synergistic eect (depending on the environmental
conditions, particularly the media) on the HHP treatments,
favoring microbial reduction (Zenker et al., 2003; Zhang et al.,
2020). According to Calvoa et al. (2018), human milk should
be discarded after a pasteurization process, when it has a to-
tal microbiological content equal to or greater than 10 CFU/
mL, in our case a higher number of bacteria was not found in
the human milk samples, after high-pressure processes and
spray drying.
Properties of HM powder
The MC of the powders ranged from 2.08 to 2.88 % (Table
5), whereas the microparticles aw showed values between
0.15 and 0.33. These results agree with the previous report of
spray-dried HM without wall materials for which the MC and
aw estimated were 1.7± 0.74 % and 0.21± 0.15, respectively
(Solís-Pacheco et al., 2019). MC and aw values lower than 5 %
and 0.6, respectively, are optimal to reduce microbiological
spoilage and prevent lipid oxidation (Sun et al., 2020).
On the other hand, the HM powder solubility was up
to 99 %, demonstrating that the macronutrients in HM did
not developed insoluble complexes with the ber, without
aecting their availability. This parameter rarely has been as-
sessed for HM, however, is widely studied in dried milk from
Table 3. Eect of spray-drying on the composition of human milk.
Tabla 3. Efecto del secado por aspersión en la composición de la leche humana.
Treatment Lactose (g/dL) Lipids (g/dL) Proteins (g/dL) Ashes (%)
Control 6.25 ± 0.75ab 2.32 ± 0.52a2.01 ± 0.63a1.50 ± 0.29a*
1 6.99 ± 0.18a2.23 ± 0.11a1.83 ± 1.00a1.32 ± 0.07a*
2 5.88 ± 0.39b* 2.18 ± 0.41a1.32 ± 0.82a1.49 ± 0.11a*
3 6.83 ± 0.39a2.06 ± 0.24a1.73 ± 0.37a1.57 ± 0.32a*
4 5.74 ± 0.13b* 1.95 ± 0.17a1.30 ± 0.30a1.34 ± 0.05a*
Values are expressed as mean ± standard deviation (n = 6). Parameters of raw human milk:
lactose 7.36 ± 0.94 g/dL; lipids 2.46 ± 0.46 g/dL; proteins 1.65 ± 0.59 g/dL; and ashes 0.26
± 0.02 %. Control: raw or unpressurized dried milk. *: Indicates a signicant dierence in
comparison with the initial value in raw milk. Values in the same column followed by die-
rent letters are signicantly dierent according to Fisher’s LSD test at p ≤ 0.05.
Table 4. Reduction of Staphylococcus aureus after the conservation process.
Tabla 4. Reducción de Staphylococcus aureus después de los tratamientos
de conservación.
Treatment HHP HHP + Spray drying
1 2.45 ± 0.53aND
2 2.40 ± 0.55aND
3 ND ND
4 ND ND
HHP: High hydrostatic pressure. Values are expressed as mean ± standard
deviation (n = 6). Initial load 6.98 ± 0.23 CFU/mL. Values in the same column
followed by dierent letters are signicantly dierent according to Fisher’s
LSD test at p ≤ 0.05. ND: the microorganism was not detected after the
treatment.
Table 5. Properties of dried human milk.
Tabla 5. Propiedades de la leche humana en polvo.
Treatment Moisture
content (%)
Water
activity
Solubility
(%)
Yield
(%)
Control 2.71 ± 0.79a0.33 ± 0.14a99.1 ± 0.04a97.02 ± 0.13a
1 2.39 ± 0.35a0.21 ± 0.04a99.0 ± 0.02a91.73 ± 1.74b
2 2.88 ± 0.49a0.20 ± 0.10ª 99.0 ± 0.05a97.02 ± 0.09a
3 2.08 ± 0.38a0.15 ± 0.05a99.4 ± 0.02a88.66 ± 1.98b
4 2.69 ± 0.89a0.18 ± 0.01a99.2 ± 0.03a92.24 ± 2.21b
Control: raw or unpressurized dried milk. Values are expressed as mean ±
standard deviation (n = 6). Values in the same column followed by dierent
letters are signicantly dierent according to Fisher’s LSD test at p ≤ 0.05.
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110
other species. For dromedary and cow’s milk, the estimated
solubility ranged from 91-95 % and 78-88 %; respectively,
at an inlet temperature of 230 °C during the drying process
(Felfoul et al., 2022). Otherwise, for dried camel milk the solu-
bility was in function of the inlet temperature, being higher
at 140 °C than at 200 °C (Habtegebriel et al., 2018). Therefore,
the dierence between this study and other reports is related
to the specie, and the drying processing parameters, due to
the milk powder obtained at a less severe processing tempe-
rature, the powder retains higher solubility of milk proteins
(Habtegebriel et al., 2018).
The yield of the spray-drying process for HM added with
soluble ber was up to 99 %. This result is higher than the
obtained for groundnut (82.72 %, inlet temperature 186 °C)
(Saha et al., 2019), dromedary and cow milk (94.99 %, inlet
temperature 230 °C, and 95.54 %, inlet temperature 230 °C,
respectively) (Felfoul et al., 2022). The yield of the spray-
drying process is strongly dependent on inlet temperature.
Generally is assumed that the use of high inlet temperatures
(180-230 °C) provides a higher yield (Felfoul et al., 2022; Saha
et al., 2019). However, this is not the only important variable
in the yield of dried milk. For camel milk, the total solids re-
covery in the cyclone increased by an increment of the inlet
temperature and airow rate, whereas decreased with an
increase in the total solids content of milk (Habtegebriel et
al., 2018). An increment in the total solids of the milk is rela-
ted to a viscosity increment, diculty feeding, and aspersion
rate resulting in droplets sticking with the drying chamber
(Habtegebriel et al., 2018; Langrish et al., 2006). Otherwise,
the high yield obtained in this research could be related to
the use of soluble ber, which, additionally, can contribute
to macronutrient maintenance. In milk, the concentration
of macronutrients in HM can be inuenced by various pro-
cesses, such as storage, freezing, thawing, and spray-drying
processes. These techniques can decrease the lipid content in
HM by up to 9.0 %, attributable to its adherence to the wall of
the dryer chamber and equipment, faults homogenization,
lipolysis, or lipid peroxidation (Cavazos-Garduño et al., 2016;
Chang et al., 2012). Otherwise, aggregation of casein micelles
produces variation in the protein concentration (Chang et al.,
2012).
CONCLUSIONS
The combination of HHP and spray-drying technique is an
alternative process that allows the maintenance of the value
of lactose, lipids, proteins, and ashes in HM. These values
were not aected by the use of each treatment alone or in
combination. HHP (300 MPa) by itself reduced the microbial
loads to undetectable levels, even for S. aureus articially
inoculated, if the process was carried out at 45 °C. However,
the use of spray-drying after the HHP at 35 °C reduced the re-
manent microorganisms. Hence, to assure the HM safety the
treatment recommendable would be HHP at 45 °C for 10 min
followed by the spray-drying process. The transformation of
an HM into a powder facilitates its storage and handling in an
HMB. The low aw and MC values are of great importance as
are related to food spoilage reduction in dried foods. Besides,
the use of soluble ber as wall material allowed us to obtain a
high yield from the spray-drying process.
ACKNOWLEDGMENTS
The authors thank Hospital Civil “Fray Antonio Alcalde” of
Guadalajara and Universidad de Guadalajara for providing
the means to carry out this work.
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