Critical Reviews in Food Science and Nutrition

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Food Processing by High Hydrostatic Pressure

M. F. San Martín , G. V. Barbosa-Cánovas & B. G. Swanson

To cite this article: M. F. San Martín , G. V. Barbosa-Cánovas & B. G. Swanson (2002) Food
Processing by High Hydrostatic Pressure, Critical Reviews in Food Science and Nutrition, 42:6,
627-645, DOI: 10.1080/20024091054274

To link to this article: http://dx.doi.org/10.1080/20024091054274

Published online: 03 Jun 2010.

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 Critical Reviews in Food Science and Nutrition, 42(6):627–645 (2002)

Food Processing by High Hydrostatic Pressure

M.F. San Martín,1 G. V. Barbosa-Cánovas,1* and B. G. Swanson2
1
Department of Biological Systems Engineering and 2Department of Food Science and Human Nutrition,
Washington State University, Pullman, WA 99164–6120

Referee: Dr. Gonul Kaletunc, The Ohio State University, Dept. Food, Ag. and Biol. Eng, 210 Ag. Eng. Building, 590 Woody Hayes
Drive, Columbus, OH 43210-1057

* Author to whom all correspondence should be addressed.

ABSTRACT: The use of high hydrostatic pressures (HHP) for food processing is finding increased application
within the food industry. One of the advantages of this technology is that because it does not use heat, sensory,
and nutritional attributes of the product remain virtually unaffected, thus yielding products with better quality than
those processed traditional methods. HHP have the ability to inactivate microorganisms as well as enzymes
responsible for shortening the life of a product. In addition to lengthening the shelf-life of food products, HHP can
modify functional properties of components such as proteins, which in turn can lead to the development of new
products. Equipment for large-scale production of HHP processed products are commercially available nowadays.
Guacamole, sliced ham, oysters, and fruit juices are some of the products currently available on the market. HHP
technology is one of the most promising nonthermal processes.

KEY WORDS: nonthermal processes, food preservation, microbial inactivation, new technologies.

I. INTRODUCTION Traditional food-processing methods have

relied on high temperatures as a way to ensure
The idea of using pressure as a process vari- prolonged shelf-life and food safety. However,
able in food processing is not new. First attempts the use of such high temperatures is commonly
date back to 1899, when Hite observed that shelf- known to cause detrimental changes on the pro-
life of milk and other food products could be cessed products. These undesirable changes af-
increased after pressurization. However, nearly fect nutritional as well as organoleptic attributes.
one century had to pass before extensive research Several vitamins degrade under heat treatments
continued in this area. as do color and flavor compounds. Texture is also
The advantages of isostatic pressing systems negatively affected, frequently vegetable tissues
have been used for cold forming of powdered soften and chemical compounds need to be added
products for ceramics, manufacture of solid to regain firmness. All these changes result in
“green” products of large and complex shapes, products that are far from similar to original fresh
powder forming of polymers and explosive pow- products. According to a recent article on the Top
ders, crystal hydrothermal synthesis, and also in Ten Trends to Watch and Work for the Next
the production of superhard materials such as Millenium, the claim for freshness in food prod-
polycrystalline stishovite1 (the high-pressure phase ucts has, since 1994, been at the top of the list of
of silica). More recently, the use of high-pressure the most desired claims by consumers. Interest-
processing has been finding application within ingly enough, this same article states that among
the food industry. the list of food issues that consumers are “very

1040-8398/02/$.50
© 2002 by CRC Press LLC
627
concerned” about is topped by E. coli and Hepa- directions, which allows solid foods to retain their
titis outbreaks.2 original shape. The pressure is held for the de-
Nowadays, consumer preferences point to- sired treatment time and then released. The ap-
ward those products that, while having longer plied pressure and the holding time will depend
shelf-life than fresh ones, have similar character- on the type of product treated and the expected
istics to the original product. This view sets a new final result, that is, generally, enzyme inactiva-
challenge for the food industry, as new processes tion requires using higher pressures than pres-
need to be developed and adopted to satisfy con- sures used for microbial inactivation.
sumer demands. As a result, novel technologies One of the advantages offered by isostatic
are currently under extensive research. Among compression over conventional thermal treatments
these nonthermal processes are the use of pulsed is the process is independent of size and geometry
electric fields (PEF), irradiation, high-intensity of the product, which are important limiting fac-
light pulses, and high hydrostatic pressures (HHP). tors in thermal processing and that frequently
Currently, the widest application of HHP pro- lead to size reduction.6
cesses within the food industry is mainly for ex- The advantages of HHP processing have let
tending the shelf-life of food products, although this process be successfully adopted by the food
as research progresses other uses are foreseen. industry, and some products thus produced may
These include solute diffusion processes3 (salt- be found in the market now. A HHP processing
ing, sugaring), assisted freezing-thawing pro- system consists basically of the pressure vessel, a
cesses,4 and modification of functional properties pressurization system, devices for temperature
of proteins and other macromolecules.5 control (heating/cooling systems), and product
It is the purpose of this article to describe the handling devices. Traditionally, pressure vessels
HHP process, to address its effects on microor- have been designed to provide treatment to small
ganisms and food components, and, finally, to volumes and at low-frequency cycles, that is, few
describe some of the equipments currently avail- compression and decompression cycles over a
able and used within the food industry, as well as given time. However, to process large volumes,
the products currently processed by this technol- the food industry requires high-frequency cycles,
ogy. that is, several compression-decompression cycles
over a given time, and, in addition, safety issues
as well as conveinence for cleaning and hygienic
II. HIGH HYDROSTATIC PRESSURE: design must be considered. Two designs are widely
THE PROCESS AND ITS USE IN THE used for the making of the pressure vessels, the
FOOD INDUSTRY monoblock and the prestressed design. Prestressed
arrays are preferred, because they allow the sys-
The HHP was adopted and adapted by the tem to leak before major failure occurs. The pat-
food industry from the isostatic pressing process ented QuintusR pressure vessel (Figure 1) consists
used from a long time ago in the ceramics indus- of a cylinder prestressed by several kilometers of
try. Variations of the process depend on specific steel wire to prevent cylinder expansion during
products, temperature, and pressure transmitting pressurization. The vessel remains prestressed
fluid (gas or water). The HHP process is quite even at maximum pressures. A replaceable steel
simple. As its name suggests, the food (liquid or liner is inserted inside the cylinder so as to cause
solid) is subjected to pressures above 100 MPa up a leak before failure, ensuring safety against ma-
to 900 MPa, with pressures used in commercial jor failures. Explosions are avoided in this way.
systems between 400 and 700 MPa. In this system top and bottom closures are kept in
The pressurization is carried out for the dura- place by a retractable prestressed frame. If the
tion of the treatment in a confined space (pressure processed product is to be in direct contact with
vessel) containing a fluid (usually water) that acts the pressure vessel, sanitary materials must be
as the pressure transmitting medium. Pressure is used in its construction, when products to be
applied isostatically, that is, equally applied in all treated are previously packaged other metal al-

628
 FIGURE 1. Patented Prestressed QuintusR
Pressure Vessel. (Courtesy of Flow Interna-
tional.)

loys may be used. The generation of pressure the simplest heating by electric heating to more
inside the vessel may be achieved by either one of complex heat-exchanging systems. Electrical heat-
three ways: ing may be used by placing heating bands outside
and around the vessel or to directly heat the pres-
1. Direct compression: The volume of the sure transmitting fluid and pump it into the vessel
treatment chamber is reduced by the action at the desired temperature. In other arrangements,
of a hydraulic pressure applied over a pis- a heating or cooling medium can be circulated
ton. through the top and bottom closures, thus acting
2. Indirect compression: In these systems an as heat exchangers. Operation temperatures will
intensifier or high-pressure pump is used to depend on the particular type of system, but the
pump the pressurizing medium directly into range can vary from –20°C to 80°C.
the vessel to reach a given pressure. This is Handling systems usually consists of baskets
the method used for the HHP application in that are filled with the packed products (such as
food processes. Examples are the system bottles or bags) that are then loaded into the HHP
used by Flow in its Isolator, and by ALSTOM unit. The baskets may be transported by convey-
in their semicontinuous system. These sys- ing belts (horizontal loading) or by cranes (top
tems also use a piston to increase the pres- loading).
sure, as in direct compression, but the dis- HHP is a batch process, although semicontinuous
placement of the piston is achieved by lines may be built by assembling three or more
pumping water with a high pressure pump pressure vessels in series. Figure 3 shows an ar-
into the other side of the piston, as shown in rangement of three pressure vessels to obtain a
Figure 2. semicontinuous process.
3. The third pressurization method, and that is
not used in the food industry so far, involves
the heating of the pressure transmitting III. WHY DOES HHP PROCESSING
medium inside the vessel to cause expan- WORK?
sion by increase in temperature.
The successful application of HHP technol-
HHP systems that allow for temperature con- ogy in the food industry results from the effects
trol are varied and the heating systems go from that high-pressure treatments cause on microor-

629
 FIGURE 2. System used for processing of bulk liquids. (Courtesy of ALSTOM.)

FIGURE 3. Semicontinuous Processing System. (Courtesy of Flow International.)

630
ganisms (spoilage and pathogens) and on some VI, and Ice VI, each of them having higher melt-
enzymes, which are the most important factors ing temperature as pressure increases, that is, the
contributing to food safety and stability. Because slope of the melting curve becomes positive. For
water is one of the main components of foods, let example, the melting temperature of ice increases
us first consider the effect that HHP has on water gradually with an increase in pressure and is 20oC
structure, then some examples of effects of HHP at 880 MPa and 30oC at 1036 MPa.8,9 The impli-
on microorganisms and in food components are cations of the behavior of water under high pres-
presented. sure are discussed later.

IV. WATER UNDER HIGH PRESSURE V. EFFECT ON BIOMEMBRANES

Water plays an important role in many bio- Biomembranes have been identified as the main
logical processes. It is considered the universal site affected by pressure.10 Biological membranes
solvent and has unique physical properties (high posses a common basic structure known as the
heat capacity, high melting and boiling tempera- fluid mosaic model proposed by Singer and
tures, high surface tension, high latent heat, reluc- Nicolson in 1972. The membrane is composed by
tance to dissolve nonpolar compounds) known as a bilayer of phospholipids with embedded func-
the anomalies of water, because they are very tional proteins that, among others, play an impor-
different from properties of similar compounds. tant role in transporting ions and other substances
The anomalous behavior of water is due to the across the membrane. It has been observed that
ability of water molecules to form hydrogen bonds lipid bilayers undergo phase transitions under pres-
that allow the formation of structured three-di- sure. Physiological phase corresponds to a liquid-
mensional networks or water clusters. A mol- crystalline phase, in which the hydrocarbon chains
ecule of water typically presents a tethraedral of the lipid bilayers are conformationally disor-
arrangement. Each water molecule is capable of dered (“melted”). Gel phases correspond to rela-
forming four hydrogen bonds, with neighboring tively ordered and more extended hydrocarbon
molecules giving place to a randomized imperfect chains. Braganza and Worcester11 identified that
space-filling network. The properties of water can multilamellar vesicles of DPPC (1,2-dipalmitoyl-
change if the volume is changed over significant 2-oleoyl-sn-glycero-3-phosphatidylcholine) on
ranges at constant temperatures. The solid state of buffer solution at 54.5°C undergo two phase tran-
water, ice, presents polymorphic forms and de- sitions from liquid crystalline to gel to an interdigi-
pending on the pressure at which freezing takes tated state, accompanied by a change in thickness
place up to nine polymorphic forms have been from 6.6 nm at 0.1 MPa to 7.5 nm at 50 MPa and
identified. Under atmospheric pressure, water crys- higher, and a further decrease to 4.8 nm at 190
tallizes at a temperature of 0°C and the hexagonal MPa. However, when other substances are present,
Ice I is the thermodynamically stable phase. This such as cholesterol, a bilayer in gel state increases
ice form has a poor packing efficiency. The appli- in thickness and decreases the cross-sectional area
cation of high pressure to water and ice results in of the hydrocarbon chains. Kato and Hayashi12
more compact structures with distorted hydro- suggest that opposite to what occurs in model
gen-bond angles and a close approach between membranes, natural membranes seem to be thick-
the nonhydrogen bonded neighbors.7 Figure 4 ened by pressure. The decrease in biomembrane
shows an approximate P-T phase diagram for fluidity as a result of a phase transition may result
water. In this diagram it is interesting to note that in breakage of the membrane and in denaturation
the melting point of Ice I decreases continuously of the membrane-bound proteins causing a func-
with increasing pressure up to a pressure of 207 tional disorder of these proteins. Some examples of
MPa and -22°C, that is, the slope of the curve is the biomembrane alterations attributed to pressure
negative up to this point. From this point on, the were reviewed by Kato and Hayashi,12 and those
polymorphic form changes to Ice III, Ice V, Ice relevant for HHP processing are the following:

631
 FIGURE 4. Phase diagram of water at high pressure.

1. Solubilization and leakage of intracellular attributes in many products, the main concern
substances from yeast cytoplasm, such as regarding its spreading is still related to food
amino acids at pressures higher than 300 safety issues. Extensive research has been done
MPa with a maximum at 600 MPa and and still continues. High pressures have the abil-
equivalent to a heat treatment of 100oC for ity to inactivate many types of vegetative cells,
5 min. but spores are resistant to pressurization up to
2. Leakage of metallic ions K+, Mn2+, Ca2+, 1200 MPa.15 Therefore, current trends also look
and Na+ at 300 MPa, and Mn2+ and Zn2+ at to the combination of HHP and some other treat-
400 MPa. ment that allows the production of safe products
3. Permeation of extracellular substances into in which spore germination could be a problem.
cells and tissues, for example, NaCl perme- Some examples of recent works to study the ef-
ation into radish tissue in 0.5% NaCl solu- fectiveness of HHP in inactivating microorgan-
tion at 400 MPa for 10 min. isms and spores are given next.

Some suggested applications of the altered mem-

brane permeability in fruit tissue are to enhance mass A. Vegetative Cells
transfer during osmotic dehydration13 or in fluidized
bed drying when combined with freezing.14 Hayert et al.16 developed a method to micro-
scopically observe directly the effect of HHP on
Saccharomyces cerevisiae cells. Immobilized cells
VI. HHP EFFECT ON MICROORGANISMS were pressurized to 250 MPa and holding time of
15 min. They observed an initial volume reduc-
Although the use of HHP has proven to be tion of 10% that increased as holding time pro-
effective in preserving nutritional and sensory gressed to a final value of 25%. Leakage of Na+,

632
Li+, and Ca2+ ions was observed. A baroprotective Rhodotorula rubra, and in some cases the same
effect at low aw values was identified when large treatment, depending on the ion, was different up
amounts of glycerol and sorbitol were used. The to 3 orders of magnitude in reduction.
reduction in viable counts was greater when ace- The sensitivity of different microorganisms to
tic acid was added, while no increased effect was HHP treatments is strongly dependent of the strain
observed for other acids such as chlorhydric, or- and under the same treatment the response may be
thophosphoric, mesaconic, pimelic, citric, lactic, greatly different. Ludwig and Schreck20 studied the
tartaric, and ascorbic. effect of different pressure treatments at different
Media composition plays an important role temperatures on the inactivation of 11 bacterial
when studying microorganism inactivation. How- strains. They identified that inactivation depends
ever, along with the protective effect that low aw on bacterial shape, with rods being more sensitive
effect on microorganisms, other factors such as than cocci, and medium sensitivity for organisms
temperature play an important role as well. with shape in between cocci and rods. No correla-
Iwahashi et al.17 showed that different strains of tion between sensitivity and gram type was ob-
Saccharomyces cerevisiae exhibited different served in this work, although other studies had
barotolerance to a pressure of 180 MPa. At low previously indicated that in general Gram-positive
temperature (4 oC) the strains showed lower organisms are more resistant to pressure than Gram-
barotolerance than at higher temperature (20oC). negative, and yeast sensitivities in between.10
The higher increase in barotolerance of strains Chilton et al.21 studied the capacity of E. coli
containing the enzyme hsp104 (heat shock pro- pressure-damaged cells to repair their outer and
tein) as opposed to those who lacked of it was inner membrane by using bile salts, NaCl, and
attributed to the fact that at higher temperatures different antibiotics to inhibit specific cellular
the activity of hsp104 was increased, thus in- processes. They concluded that damaged cells are
creasing barotolerance. In another work for the able to repair the outer membrane even when
mutant CWG8 of Saccharomyces cerevisiae, a antibiotics are present, while repair of the inner
mutant that showed the highest tolerance against membrane requires the action of many cellular
temperature, pressure, and oxidative stress among processes. They also studied damage caused to
tested mutants, Fujii et al.18 concluded that the nuclear material. They hypothesized a possible
level of heat shock protein in this strain was insuf- aggregation of ribosomes or cytoplasmic units,
ficient to provide barotolerance, but that trehalose and by electrophoresis they detected cleavage of
content and membrane fluidity were the major DNA strands and proved it to be enzymatically
factors contributing to barotolerance. controlled. They suggest that cleavage of genetic
The nature of the ions present in a media material may be another cause for cell death.
along with the kind of microorganism can modify The ability of certain strains to develop
the barotolerance of the organism as shown by barotolerance was showed by Hauben et al.22
Michiels et al.19 They studied the effect of differ- E. coli cells were subjected to successive pressur-
ent ions (K+, Li+, Na+, Ca2+, Mn2+, Mg2+, NH4+) on ization cycles, with regrowth of the survivors
the barotolerance of a Rhodotorula rubra strain. after each cycle. They showed that significant
At 160 MPa, Li +, and Na + ions increased barotolerance was developed. For the parent strain
barotolerance significantly at low concentrations a HHP treatment of 220 MPa resulted in 10% of
(<10 mM). The increased barotolerance did not surviving cells, whereas at this same pressure
occur in other studied yeasts, except for 50% of the barotolerant mutant survived, and in-
Schizosaccharomyces pombe, which showed in- activation increased marginally from 50% survi-
creased barotolerance dependent on solute con- vors at 220 MPa to 0.1% survivors at 800 MPa.
centration, but was not ion specific. The effect They concluded that accumulation of pressure-
conferred by the presence of Na+ ions was linked resistant mutants on processing equipment may
to pH homeostasis and related to Na+-specific put at risk the safety of HHP processed foods,
transport processes in the membrane. Na + because these barotolerant mutants were obtained
baroprotective effect was only observed in after only 18 pressurization cycles.

633
 The age of the culture influences the inactiva- In regards to the use of bacteriocins in com-
tion rate, with stationary cells being more resis- bination with HHP treatment, these compounds
tant than exponential phase cells as demonstrated may exhibit synergistic effects as shown by Ponce
by Van Almsick and Ludwig.23 These researchers et al.28 They studied the effect of nisin on Listeria
also showed that inactivation of vegetative anaero- innocua and E. coli on liquid whole egg and
bic bacteria did not significantly differ from aero- found that the addition of nisin significantly in-
bic bacteria inactivation. creased the inactivation effect of HHP pressure
Patterson et al.24 demonstrated the medium treatment on both microorganisms. The amount
effect on the inactivation of vegetative pathoghens of nisin added had an effect on Listeria innocua,
of Yersinia enterocolitica, Salmonella typhimurium, but not in E. coli. Garcia-Graells et al.26 reported
Listeria monocytogenes, Salmonella enteriditis, that the effect of nisin and lysozyme on E. coli
Escherichia coli O157:H7, and Staphylococcus can be enhanced by cyclic pressure treatments.
aureus. They confirmed that sensitivity of every Erkmen and Karatas, 29 studied the effect of
microorganism was different and dependent on the HHP on Staphylococcus aureus innoculated in
medium used. For example, when treated in a pasteurized whole cow’s milk finding a reduction
phosphate-buffered saline, S. aureus was the most level of 2,3 and 8 log cycles for treatments at 200,
pressure resistant. However, when studied using 250, and 300 MPa, respectively, for 10 min.
UHT milk, E. coli O157:H7 was the most resis- In addition to inactivation, high pressure also
tant. They also demonstrated that variability in induces morphological changes. These are classi-
resistance was also dependent on the species and fied as reversible changes such as elongation and
on strain among species. When comparing the cessation of motion due to structural changes, and
type of media, UHT milk was more protecting to irreversible changes such as separation of the cell
L. monocytogenes, E. coli O157:H7, and S. aureus wall from the cell membrane as well as a decrease
than poultry meat. They also observed sublethal in the number of ribosomes.10
injury at pressures lower than that required for Some of the factors that affect inactivation
cell death and suggested that this effect could be are shown in Table 1. Table 2 shows some ex-
used advantageously in combination with other amples of results for HHP inactivation on several
treatments such as mild heating. The protective microorganisms.
effect provided by the type of media was also
confirmed by Gervilla et al.25 when studying the
inactivation effect of HHP on Listeria innocua B. Spores
inoculated in Ewe’s milk. Significant differences
between inactivation achieved at 200, 300, and One of the major drawbacks of HHP is the
350 MPa were found, while inactivation observed inability to inactivate spores by pressure alone.
at 400, 450, and 500 MPa were not significantly Spores have shown to be very pressure resistant,
different. They suggested a baroprotective effect capable to survive pressures up to 1200 MPa.42,43,44
provided by fat. Similar protecting effects by the However, it has been demonstrated that low-pres-
medium have been reported by Garcia-Graells et sure treatments (60 to 100 MPa) can induce spore
al.26 on E. coli in milk. germination and that this process is temperature
The growth conditions of the culture also dependent, with higher temperatures leading to
play an important role in the response of the higher percentages of germinated spores. The
microorganisms to HHP treatment. Mc Clements mode of action has been suggested to be by
et al. 27 demonstrated that cultures of electrostriction or an increase in ionization of
L. monocytogenes strains NCTC 11994 and Scott some spore components that still remains specu-
A in stationary phase showed higher inactiva- lative. Electrostriction involves a decrease in the
tion rate when grown at 30oC than those grown system volume caused by the orientation that water
at 8oC. However, for B. cereus stationary-phase molecules experience due to the high electric field
cells grown at 30oC were more resistant than of the ion, causing a local collapse of the bulk
those grown at 8oC. water structure. Electrostriction causes a com-

634
 TABLE 1
Factors Contributing to Inactivation of
Microorganisms

pression of adjacent dipoles and is the result of five cycles of oscillatory pressurization at 689
ion-water interactions that depends on the radius MPa, 60oC and aw 0.98, whereas less than one
of the ion and on its charge. Electrostriction would cycle reduction was observed when aw was 0.94.
allow hydration of the core, thus beginning the Under same pressure and water activities but with
germination process.42 The germinated spores can temperature of 21oC no significant reduction was
then be inactivated by treatments at higher pres- observed.
sures. Moreover, it has been demonstrated that Nakayama et al.44 studied the pressure resis-
the pressure used to induce germination will af- tance of six bacillus strains and compared it to their
fect the sensitivity of germinated spores to subse- heat resistance. Spores from B. stearothermophilus,
quent stresses. Wuytack et al.45 observed that B. subtilis, and B. licheniformis could not be inac-
Bacillus subtilis spores germinated at 100 MPa tivated by pressure treatments of 981 MPa for 40
were more sensitive to pressure, UV light, and min or 588 min for 120 min. Temperatures used
hydrogen peroxide than spores germinated at 500 were 5 and 10oC. No correlation between pressure
MPa. It must be noted, however, that the times and heat resistance was observed by these research-
required to achieve spore germination were long, ers. On the other hand, Hayakawa et al.46 reported
in the order of 30 min. that 4 log cycle reduction of B. stearothermophilus
The manner in which pressure is applied, spores was obtained at 70oC after six pressuriza-
namely, continuously or by cycles, has a signifi- tion cycles at 400 MPa, and six log reduction at
cant effect on the inactivation achieved, with pres- 600 MPa and 70oC.
sure treatment applied as cycles being, by far, Raso et al.48 reported that sporulation tem-
more effective.46,47 Palou et al.47 observed that perature is another factor that influences the inac-
inactivation of Bysocchlamys nivea ascospores in tivation of Bacillus cereus germinated spores
fruit juice concentrate depended on temperature, exposed to high pressure. Higher sporulation tem-
water activity, and nature of the pressure treat- perature (37oC) resulted in higher germination
ment (oscillatory or continuous). For example, up and inactivation rate when compared with
to four log cycles could be reduced after three or B. cereus sporulated at lower temperature (20oC).

635
 TABLE 2
Examples of Inactivation of Different Microorganisms at Various HHP Treatments

636
 Reddy et al.49 reported that no reduction in were more effective. Treatments at 500 MPa for
Clostridium botulinum Type E spore content was 10 min, 700 MPa for 1 min, and 900 MPa for
observed at 35oC and pressures from 414 to 827 1 and 10 min yielded a product with cloud stable
MPa for 5 min, whereas when temperature in- for up to 90 days. All treatments yielded micro-
creased to 55oC up to 5 log cycle reduction was biologically stable products. According to their
observed. results, HHP treatment of orange juice is a viable
Zook et al.50 obtained D and z values for alternative to conventional thermal treatments.
inactivation of S. cerevisiae ascospores in orange Some enzymes have been reported to increase
juice and in a model juice, and, by comparison their activity after pressurization, such as
between these values, concluded that juice com- polyphenoloxidase (PPO) at 400 MPa in mush-
ponents did not have any effect on the lethality of rooms.55 The effects of HHP on enzymes are
the ascospores. dependent on type of enzyme, source, substrate,
Inactivation of spores from Clostridium pressure, time, and temperature. In some cases
sporogenes could not be inactivated by pressure combination of HHP with certain pretreatments
alone, and for none of the treatments tested (400 may be required to achieve enzymatic inactiva-
MPa at 60oC for 30 min, or 80oC for 10 min tion. Palou et al.53 showed that residual PPO ac-
followed by 400 MPa for 30 min) the reduction tivity in banana puree after HHP treatment (517
was greater than 3 log cycles.51 and 689 MPa for 10 min) can be decreased when
The combination of HHP with other treat- a blanching pretreatment is given. They observed
ments has been suggested as a way of reducing that residual activity <5% could be obtained if
spore number. The use of a high-pressure treat- blanching with saturated steam for 7 min fol-
ment (689 MPa at 80oC for 20 min) of Clostridium lowed by HHP treatment at 689 MPa for 10 min
sporogenes spores in chicken breast combined was applied. However, this amount of residual
with irradiation by an electron beam lowered the activity was enough to initiate browning after
irradiation D value from 4.1 to 2 kGy.52 6 days of storage at 25oC. Hence, a suitable com-
From all the previous works it can be seen bination of treatments must be studied to obtain
that spores are highly resistant to pressure, and stable products. pH plays an important role in
that in some cases the temperatures required to PPO inactivation. Eshtiaghi and Knorr56 could
significantly decrease spore population are very completely inactivate PPO at 400 MPa within
high as well. Therefore, research in this area is 15 min using 0.5% citric acid solutions.
still required if HHP is to be used in the produc- Gomes and Ledward57 studied the inactiva-
tion of low-acid shelf-stable foods. tion of some polyphenoloxidases in potato, mush-
room, apple, mushroom extract, and in the puri-
fied enzyme. They observed that in mushrooms
VIII. HHP EFFECT ON PROTEINS browning rate is in fact increased after a pressure
treatment of 400 MPa. In potatoes and apples
A. Enzymes browning occurred at pressures ranging from 200
to 600 MPa, while at 800 MPa complete inactiva-
The response of enzymes subjected to HHP tion of the enzyme was achieved. Pressure treat-
treatments is varied and depends on the origin of ment of the purified enzyme did not cause an
the enzyme, nature of the substrates, pressure, enhancement in activity. They concluded that
temperature, and time of processing.53 The stabil- activity enhancement may be due to changes in
ity of cloud, whose loss is caused by the action of interactions between extract constituents or from
the enzyme pectinesterase, in orange juice treated the release of membrane-bound enzymes. The
by HHP was studied by Goodner et al.54 They HHP resistance for polyphenoloxidase was also
reported that pressures greater than 500 MPa and demonstrated by Weemaes et al.58 who showed
times longer than 1 min were required to increase that pressures of 800 MPa were needed to inacti-
cloud stability when compared with untreated vate polyphenoloxydase from mushrooms. They
samples, and that higher pressures for longer times also demonstrated that HHP PPO inactivation from

637
different fruits follow first-order kinetics. PPO C. Cold Denaturation of Proteins
from apple, grape, avocado, and pear began show-
ing reduction at 600, 700, 800, and 900 MPa at The anomalous properties of water under pres-
25oC, respectively, whereas for plum reduction sure allow the liquid to undergo significant super-
only began to be noticed at 900 MPa and 50oC. cooling at elevated pressures. This fact has been
Increase enzymatic activity was also observed used to study the denaturation of proteins at sub-
by Gomes et al.59 for α− and β-amylases from malt zero temperatures without freezing. Cold dena-
barley when 10% wheat or barley flour slurries turation of proteins attracted scientific attention
were subjected to pressures between 400 and 600 some years ago. According to Le Chatelier’s Prin-
MPa. Within this range enzymatic activity was ciple, a decrease in temperature in a system should
increased, but after 20 min at 600 MPa activity lead to a decrease in enthalpy and entropy, that is,
decreased, suggesting that inactivation is pressure increasing order. Hence, because proteins’ high-
and time dependent. At higher pressures (i.e., 700 est ordered state was supposed to be their native
to 800 MPa) activity was significantly decreased. state, a temperature decrease could not be ex-
These researchers suggest that within the range pected to lead to drastic conformational changes.
400 to 600 MPa starch granules gelatinize, and the However, experimentally it was found that globu-
active site of the enzyme is modified in a way such lar proteins undergo a change of protein structure
that hydrolysis is favored, whereas at 700 to 800 and a disruption at very low temperatures.65 Jonas7
MPa even though gelatinization of starch granules studied the structure of pressure denatured and
occurs, the enzyme looses its activity. pressure-assisted cold-denatured (cold denatur-
ation at constant high pressure) ribonuclease A
and compared with the heat denatured state, and
B. Protein Gelation concluded that pressure and pressure-assisted cold-
denatured enzyme have “more” secondary struc-
Gel formation by proteins was observed as ture than heat-denatured enzyme, and point out
early as 1914 on the coagulation of albumen by that more studies with other proteins are neces-
pressure.60 High pressures can induce gelation at sary to determine whether the observed effect can
low temperatures, resulting in gels with proper- be generalized. High pressures can assist in the
ties different from gels obtain with heat. Chung et cold denaturation of carboxypeptidase Y as
al.61 reported that surimi gels from Pacific whit- demonstratec by Kunugi et al.66 Figure 5 shows a
ing obtained by HHP were less opaque than tra- schematic diagram of protein denaturation. C and
ditionally heat set gels and had higher stress and H define the points at which cold and heat dena-
strain values than those of heat set gels. Ashie and turation occur at atmospheric pressure. Tempera-
Lanier62 reported that surimi gels form by pres- ture and pressure combinations that led to protein
sure were less opaque and more glossy than cooked denaturation states are delimited by an ellipsoidal
gels, but cooked gels were stronger. In this same curve. A phase diagram of water is represented in
study, pastes of turkey did not formed gels by this figure as well. It can be seen that at atmo-
pressure treatments alone, but the addition of spheric pressure the formation of ice due to the
microbial transglatuminase resulted in gelation low temperatures does not allow to study cold
under pressure, but the strength of these gels was denaturation, whereas if increasing the pressure
lower than heat formed gels. to cool water to subzero temperatures ice is not
Kumeno63 reported that pressure-induced gels formed and cold denaturation occurs.
from freeze-concentrated milk had high white-
ness, high brightness, and fresh cream-like flavor,
and suggested its use for high-quality cold des- IX. HYDROSOLUBLE VITAMINS AS
serts manufacture. AFFECTED BY HHP
Treatment of albumen by HHP renders a mi-
crobiological stable product, retains its functional One of the advantages constantly mentioned
properties, and increases protein digestibility.64 when speaking about HHP is the retention of

638
 FIGURE 5. Schematic diagram of protein denaturation.

original nutritional quality of the product. High degree of vitamin C retention is independent of
pressure affects the interaction of components by the pressure applied (200, 400, or 600 MPa, for
changing the distance between them. Because the 30 min), and concluded that HHP treatment con-
length of a covalent bond is yet limited by repul- tributes to preservation of nutritional quality of
sions at short interatomic distances, known as foods while extending their shelf-life.
Born repulsions, interactions where a volume
decrease is possible, such as hydrogen bonds, are
more likely to be affected by high pressures.9 It X. EFFECT OF HHP ON VOLATILE
has been hypothesized that because HHP does not PROFILES
affect covalent bonds, small molecules such as
vitamins, color, and flavor compounds will re- The use of high pressures (800 MPa) have
main unaffected after the treatment. been shown to affect the volatile profile of certain
Sancho et al.67 studied the effect of a HHP products. Lambert et al.68 studied the effect of
treatment on the retention of the hydrosoluble HHP treatments on the volatiles profile of a straw-
vitamins B1, B6, and C after HHP treatment in a berry coulis and compared with to samples of
multivitamin model system in egg yolk and in fresh and sterilized (20 min at 120oC) coulis.
strawberry “coulis” and compared it to the reten- They reported that pressure treatment of 200 and
tion achieved after a high temperature pasteuriza- 500 MPa at 20oC for 20 min do not significantly
tion (20 s at 76oC) and sterilization (20 min at affect the volatiles profile, but treatments at 800-
120oC). They found that HHP does not have a MPa result in a different profile, as analyzed by a
significant effect on the retention of either B1 or gas chromatography-mass spectrometry technique.
B6 vitamins when compared with thermal treat- The 800 MPa treatment promotes the formation
ments, and that retention was always greater than of two new compounds that do not appear at
99%. On the other hand, the retention of vitamin lower pressures. Sterilized product also showed a
C was found to be significantly different depend- significantly different profile, and compounds such
ing on the process. In strawberry coulis, retention as vanillin and an unknown compound (referred
of vitamin C was not significantly different be- to as unknown “cooked”) formed due to the heat-
tween pasteurization and HHP processing (91.52 ing effect and that is typically present in heated
and 88.68%, respectively), but was significantly products were identified. These compounds were,
different when samples were sterilized (67.11% in HHP-treated samples, either absent or on such
retention). By studying the degradation of vita- a small concentration that were not detected.
min C over time, they found that the rate of Because this work was carried out only for ana-
change is almost identical to degradation observed lytical purposes, it is not possible to determine
for the untreated product. They also found that the whether the changes promoted by a high-pressure

639
treatment of 800 MPa would have a significant teristic behavior of phase transitions of water under
sensory impact. The important fact is that the high pressure can be used to freeze or thaw prod-
treatments at lower pressures, namely, 200 and ucts that will have a better quality. Some terms
500 MPa, did not change the profile significantly. used when dealing with phase transitions are de-
Tropical fruits that contain high concentration fined by Knorr70 as:
of volatiles and are susceptible to flavor deteriora-
tion by heat treatments are good candidates for HHP • Pressure-assisted: means a phase transition at
processing. Yen and Lin69 reported that flavor of constant pressure. In Figure 4 this behavior is
guava juice was changed after heat treatment at illustrated by a vertical line, and it is obtained
95oC for 5 min. No new compounds were identified by increasing the pressure and then decreasing
after heat treatment, but most of the volatiles were (freezing) or increasing (thawing) the tempera-
lost. When high pressure treated, volatile compounds ture and finally decreasing the pressure.
content was similar to content of fresh guava juice. • Pressure-shift: means phase transition due to
The flavor remained stable for up to 30 days when a pressure change. In this case the pressure is
storage temperature was 4oC, on the other hand, increased, the temperature decreased without
when stored at 25oC the volatile profile changed changing phase, and then decreasing the pres-
significantly. This effect was attributed to the effect sure, with change of phase occurring at a lower
of enzymes that were not completely inactivated by pressure.
the HP treatment (600 MPa, 15 min). • Pressure-induced: means phase transition ini-
tiated with pressure change.

XI. OTHER EFFECTS OF HHP Among the advantages of using pressure for
freezing and thawing are a 2/3 reduction of the
A. Food Freezing and Thawing time needed for thawing frozen meat, smaller ice
crystals formation, less damage of cell structure,
Freezing and thawing of foods can be modi- and less browning in potato,70 resulting in prod-
fied advantageously by using HHP. The charac- ucts of better quality.

FIGURE 6. Fresher under pressure system. (Cour-

tesy of Flow International.)

640
B. Meat Tenderization and flavor development in many cooked products.
The reaction occurs in two or three steps, first the
Application of HHP on meat causes tender- formation of amadori compounds that are then
ization by changes on the myofibrillar pro- transformed into melanoidins and volatiles. They
teins.38,71,72 From an SEM study, Ueno et al.73 reported that when the reaction occurred under
observed that high pressure may have a different high pressure, the relative concentration of 5-me-
effect on the membrane structure of the intramus- thyl-4-hydroxy-3(2H)-furanone,1 (the main vola-
cular connective tissue than aging, but they sug- tile constituent from xylose and lysine reaction)
gested that further research is required to clarify decreases as pressure increased (0.1, 100, 200,
the origin of this difference. 400, and 600 MPa), and at the highest pressure
almost none was formed. A similar behavior was
observed for the formation of other volatiles. In
C. Reduction in Patulin Content in Apple general, the pressure effect was observed on the
Juice amount of product formed rather than on the rate of
formation. They concluded that in products pro-
Chemical reactions that occur as a consequence cessed under high pressure at temperatures suffi-
of HHP treatment can sometimes be used advanta- ciently high so that Maillard reaction occurs, the
geously. One example of this is the decrease on development of cooked flavors may be diminished.
patulin content (a toxin produced by some species However, no reports of cooking or processing un-
of Aspergillus, Penicillium, and Bysochlamys) ob- der such high pressures is available so far.
served by Bruna et al.74 in apple concentrate and In a glucose-lysine model system, Hill et al.76
apple juice. These authors point out that thermal observed that incubation under 600 MPa resulted
treatment does not result in a significant reduction in the same degree of browning as the system
of patulin content, whereas HHP treatment of 300, incubated at atmospheric temperature; however,
500, and 800 MPa at ambient temperature for 60 the yield of volatile compounds was significantly
min result in a decrease of original patulin content suppressed by high pressure. This effect was at-
of 16, 20, and 23% in apple concentrate, and 42, tributed to reaction of these compounds by aldol
53, and 62% in apple juice, respectively. They also condensations, which is a path whose rate has
found that by increasing the temperature (to 50oC) been observed to increase under high pressure.
the reduction rate can be increased. It was found
that osmotic pressure plays and important role,
because reduction was higher in the juice than in E. Milk Products
the concentrate. A similar baroprotective effect has
been identified to occur when studying the inacti- The use of HHP on milk prior to cheese mak-
vation of Byssochlamys nivea ascospores in low ing causes a decrease of rennet coagulation time,
water activity environments.47 higher wet-weight yield, as well as increased gel
rigidity77,78 by increased exposure of hydrophobic
sites and increased number of network strands.79
D. Maillard Reaction Under High Messens et al.80 demonstrated that brining of
Pressure Gouda cheese at pressures higher than 200 MPa
accelerates the hydrolysis of κ-casein, and that it
Bristow and Isaacs75 studied the chemical increases with pressure.
changes on a Maillard reaction carried out under
high-pressure treatment on a model system con-
taining xylose and lysine monohydrochloride. XII. COMMERCIALLY AVAILABLE
Maillard reaction, also known as nonenzymatic EQUIPMENT AND FOOD PRODUCTS
browning, occurs between a sugar and an amino PROCESSED BY HHP
group at temperatures higher than 80oC, leading to
the formation of brown polymers (melanoidins) First HHP processed foods were introduced
and different volatiles responsible for browning for the first time to the Japanese market in 1990,

641
and slowly HHP processed foods are being gradu- shown an interest in HHP. The latter for the pro-
ally introduced in other countries. In the U.S. duction of carrot and soy-based juices.81
HP-processed guacamole is currently on the market. A new, interesting application of HHP is for
Guacamole is processed in semicontinuous and in processing oysters. HHP has been shown to inac-
batch process using a Fresher Under PressureR Sys- tivate some species of Vibrio, while keeping the
tem. Skin and stone are manually removed from the flavor and texture unaltered, and an additional
avocado pulp. The guacamole is either packed in benefit is that oysters shuck under pressure.82
plastic pouches for batch processing in a QuintusR Another available industrial equipment is manu-
vessel or pumped to the isolators for continuous factured by UHDE, Uhde Hochdrucktechnik GmbH
processing; in this case the product is packed after in Germany and represented by HPR Inc. in the
HP treatment. Figure 6 shows Flow International’s United States. The process system is modular and
Fresher Under Pressure System. can be expanded. This equipment is a batch unit
In Spain, sliced ham pasteurized with HHP with a unique design. Top and bottom closures are
by Espuña is currently produced using an kept in place by a static frame; the pressure vessel
ALSTOM’s HYPERBAR horizontal unit. Ham is can be tilted to allow for top loading and bottom
sliced, packed in pouches, and HP treated. Shelf- unloading reducing the loading-unloading time.
life of sliced ham was extended from 3 weeks
without HP treatment to 8 weeks after pressuriza-
tion. Figure 7 shows Alstom’s Hyperbar equip- FINAL REMARKS
ment at Espuña’s ham processing plant.
In France, orange and grapefruit juices are HHP processing is finding increasing appli-
processed using HHP by the French company cation within the food industry. Although still an
ULTI also using a HYPERBAR equipment. Juice expensive process, it is of significance in the pro-
is extracted, packed in flexible bottles, and sub- duction of high-added-value products and prod-
jected to HP treatment. The shelf-life of this prod- ucts that are degraded by conventional heat treat-
uct is extended from 5 to 8 days for nonprocessed ments. As research continues, more applications
product to about 3 weeks for the pressurized juice for this technology are foreseen.
under refrigerated storage. Another French com-
pany, Pampryl, a subsidiary of Pernord-Ricard, is
processing freshly squeezed orange juice. A Japa- REFERENCES
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FIGURE 7. Hyperbar for ham processing. (Courtesy of ALSTOM.)

642
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