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Research Note
University of Leeds, Procter Department of Food Science, Leeds LS2 9JT (U.K.)
(Received April 5, 2001; accepted July 19, 2001)
The effect of high hydrostatic pressure on ester formation and hydrolysis was studied. Six esters and the corresponding carboxylic
acids and alcohols were subjected to high-pressure treatments of 400 and 800 MPa under three different pH conditions (namely,
buffer solutions of pH 4, 6 and 8). The selected compounds were dissolved into buffer solutions, subjected to the pressure treatment
and then extracted using dichloromethane. The analysis and quantification were carried out by gas chromatography with flame
ionization as detector. High pressure appeared to have no effect on ester formation or hydrolysis under the investigated conditions. In
all cases, a small decrease at the levels of carboxylic acids and esters was observed without any evidence of further reaction. This
decrease, referred to as decomposition, depended on pressure and pH conditions. Ester decomposition was minimised when a high-
pressure treatment of 400 MPa in basic conditions (pH 8) was applied. Carboxylic acid decomposition was minimal in basic
conditions and it was independent of the pressure applied.
Introduction odours and are responsible for the flavour and fragrance
of many fruits, flowers and artificial flavourings.
High hydrostatic pressure (HHP) technology is a novel Generally, the delicacy of natural flavours and fra-
nonthermal food processing technology where foods are grances is due to complex mixtures, for instance, more
subjected to HHP, generally in the range of 100–600 MPa, than 100 substances contribute to the flavour of ripe
at or around room temperature (Mertens, 1995). strawberries (Zabetakis & Holden, 1997). On the other
In general, nowadays, food consumers have a high hand, cheap artificial flavourings are often single
expectation of food quality. There is a strong demand compounds or at the most simple mixtures of esters
for more ‘natural’ minimally processed, additive free and ketones.
foods and the reported beneficial and unique effects of Among the most common esters contributing to flavour
high pressure appear to offer particular potential in and aroma are: pentyl acetate which resembles the
these respects (Johnston, 1994). odour of bananas, octyl acetate that of oranges, ethyl
Esters are among the most widespread of all naturally butanoate that of pineapples, pentyl butanoate that of
occurring compounds. They are moderately polar and apricot and isopentyl valerate that of apples (McMurry,
thus insoluble in water with dipole moments in the 1994). Our particular interest lies towards: ethyl and
1.5–2.0 D range. Dipole–dipole interactions contribute methyl acetate, ethyl and methyl butanoate and ethyl
to the intermolecular attractive forces that cause esters and methyl hexanoate which contribute to the estery
to have boiling points slightly higher than hydrocarbons and green notes of odour in strawberries. We are also
of similar molecular weight. Because they lack hydroxyl interested in their formation and hydrolysis to car-
groups, they cannot form hydrogen bonds with boxylic acids and alcohols (Zabetakis & Holden, 1997;
each other; consequently, esters have lower boiling Sumitani et al., 1994; Zabetakis et al., 2000; Larsen
points than alcohols of comparable molecular weight et al., 1992).
(Yamamoto et al., 1991). On a study on the effect of HHP on strawberry fla-
Many esters, especially the volatile ones of low vour, volatile esters were not identified (Zabetakis
molecular weight, have characteristically pleasant et al., 2000a). It was suggested that esters were
hydrolysed to the respective acids and alcohols during
HHP treatment. The absence of esters was explained
*To whom correspondence should be addressed. Stratigou Makriyanni
6, Nea Filadelfia, Athens, GR-143 42 Greece. Tel./fax: +30 10
on the basis of a possible hydrolytic effect that HHP
2510589; E-mail: dryz@subdimension.com causes (Tauscher, 1995). In this work, we report our
0023-6438/02/$35.00 doi:10.1006/fstl.2001.0863
r 2002 Elsevier Science Ltd. All rights reserved. All articles available online at http://www.idealibrary.com on
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condition for methyl butanoate is a high-pressure butanoic acid in butanoic acid–ethanol solutions is a
treatment of 400 MPa at pH 4. In the case of ethyl high-pressure treatment of 400 MPa in pH 8. For the
hexanoate, the percent decomposition varied between mixture of butanoic acid–methanol, the percent decom-
0.1 and 12%. The highest degree of decomposition was position of the butanoic acid varied between 0.5 and
observed in pH 4 and 800 MPa conditions, while the 10%. The highest degree of acid decomposition was
lowest appears to be at pH 8 when a pressure of observed at pH 4 and when the sample was subjected to
800 MPa was, respectively, applied. As a result, the 800 MPa pressure, while the lowest was at pH 8 under
optimum conditions for ethyl hexanoate is a high- 400 MPa pressure treatment. As a result, the optimum
pressure treatment of 800 MPa at pH 8. For methyl conditions for butanoic acid in butanoic acid–methanol
hexanoate, the percent decomposition varied between solutions is a high-pressure treatment of 400 MPa in pH
0.1% and 18.5%. The highest degree of decomposition 8. In the case of hexanoic acid–ethanol, the percent
was observed in pH 8 and 800 MPa conditions, while decomposition of the hexanoic acid varied between 2
the lowest appears to be at pH 8 when a pressure and 14%. The highest degree of acid decomposition was
of 400 MPa was applied. As a result, the optimum observed at pH 4 when the sample was subjected to a
condition for methyl hexanoate is a high-pressure 400 MPa pressure, while the lowest was found at pH 4
treatment of 400 MPa at pH 8. These results are under 800 MPa pressure treatment. As a result, the
summarised in Table 1. optimum condition for hexanoic acid in hexanoic acid–
methanol solutions is a high-pressure treatment of
400 MPa in pH 8. For the mixture of hexanoic acid–
Effect of HHP on acids and alcohols methanol, the percent decomposition of the acid varied
As far as carboxylic acids and alcohols are concerned, between 0.1 and 19%. The highest degree of acid
there was no sign of ester formation at all. It was found decomposition was observed at pH 4 and when the
that there was no ester present after the samples were sample was subjected to a 400 MPa pressure, while the
subjected to high-pressure treatment, while there was lowest was at pH 6 under 400 MPa pressure treatment.
only a small decrease in the carboxylic acid concentra- As a result, the optimum condition for hexanoic acid in
tion further described as decomposition. hexanoic acid–ethanol solutions is a high-pressure
In the case of acetic acid–ethanol, the percent decom- treatment of 400 MPa in pH 6. In general, the
position of the acetic acid varied between 0.1 and 8%. decomposition of hexanoic acid was very low when the
The highest degree of decomposition was observed in acid was subjected to various pressures under different
pH 4 and 800 MPa conditions, while the lowest appears pH conditions. These results are shown in Table 2.
to be at pH 8 when a pressure of 800 MPa was applied. A pattern describing the decomposition of the esters and
As a result, the optimum conditions, for acetic acid in acids cannot be established. In fact, there seems to be an
acetic acid–ethanol solutions is a high-pressure treat- antagonistic effect between two factors when pressure is
ment of 800 MPa at pH 8. For the mixture of acetic applied. In the case of high-pressure treatment, a volume
acid–methanol, the percentage decomposition of the decrease is expected to occur when pressure increases.
acetic acid varied between 0.5 and 6%. The highest But as far as high-pressure treatment on the overall
degree of decomposition was observed in pH 4 and esterification reaction is concerned, the effect cannot be
800 MPa, while the lowest in pH 6 and 400 MPa easily predicted.
conditions. As a result, the optimum conditions for In the case of ester solutions, only a small decrease in the
acetic acid in acetic acid–methanol solutions is a high- ester concentration was observed without any evidence of
pressure treatment of 400 MPa in pH 6. In the case of further reaction. There was no carboxylic acid formation
butanoic acid–ethanol, the percent decomposition of the that would indicate ester hydrolysis. Esters did not
butanoic acid varied between 0.5 and 5.5%. The highest hydrolyse when pressure was applied because when
degree of acid decomposition was observed at pH 4 hydrolysis occurs there is a volume increase. According
when the sample was subjected to 800 MPa pressure, to the principle of Le Chatelier, when pressure increases a
while the lowest was at pH 8 under 400 MPa pressure decrease in volume is expected (McMurry, 1994). Ester
treatment. As a result, the optimum conditions for hydrolysis leads to the formation of a carboxylic acid and
Table 1 The effect of HHP on ethyl acetate, methyl acetate, ethyl butanoate, methyl butanoate, ethyl hexanoate
and methyl hexanoate expressed as percent decomposition of the ester
% Decompositiona
pH 4 pH 6 pH 8
400 MPa 800 MPa 400 MPa 800 MPa 400 MPa 800 MPa
Ethyl acetate 5.270.31 7.870.42 10.170.55 8.270.44 2.170.11 13.770.56
Methyl acetate 7.870.39 10.170.45 8.270.48 10.270.51 6.470.48 5.870.38
Ethyl butanoate 0.170.01 3.170.11 0.370.02 7.570.33 3.470.23 4.370.28
Methyl butanoate 0.170.01 14.170.43 18.070.45 4.170.11 4.470.23 6.270.38
Ethyl hexanoate 1.670.12 11.870.34 4.470.29 5.970.44 4.370.22 0.170.01
Methyl hexanoate 0.970.03 2.870.11 10.170.31 6.270.29 0.170.01 18.570.59
a
Values are means and standard deviations for N=3.
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Table 2 The effect of HHP on mixtures of acetic acid/ethanol, acetic acid/methanol, butanoic acid/ethanol,
butanoic acid/methanol, hexanoic acid/ethanol and hexanoic acid/methanol expressed as percent decomposition of
the acid
% Decompositiona
pH 4 pH 6 pH 8
400 MPa 800 MPa 400 MPa 800 MPa 400 MPa 800 MPa
Acetic acid/ethanol 2.270.11 8.270.33 4.370.28 2.970.14 0.4170.05 0.170.01
Acetic acid/methanol 1.270.04 5.870.29 0.570.04 0.770.05 5.870.23 0.670.05
Butanoic acid/ethanol 2.170.14 5.570.23 1.270.14 2.370.19 0.570.05 0.970.07
Butanoicacid/methanol 4.170.24 10.170.43 2.170.14 8.170.44 0.570.02 7.870.38
Hexanoic acid/ethanol 14.170.44 2.870.34 6.870.44 5.570.33 2.070.11 7.070.22
Hexanoic acid/methanol 19.170.55 0.570.08 0.170.02 0.970.06 0.370.04 1.570.23
a
Values are means and standard deviations for N=3.
an alcohol, which causes a volume increase. Thus, ester condition under which the degree of decomposition
hydrolysis is not favoured by pressure. minimised and thus appeared to be more stable was a
On the other hand, solvolytic reactions are enhanced by HHP treatment of 400 MPa at pH 8. In the case of
pressure. Solvolytic reactions are nucleophilic substitu- carboxylic acids, high pressure appeared to have no
tions by the solvent, which, in this case, is water significant effect on the degree of decomposition.
(McMurry, 1994). As a result, the solvent nucleophili- Minimal loss was observed at pH 8, independent of
cally attacks the ester and enhances its hydrolysis. the pressure applied. Acids appeared to be stable at a
However, the results that we obtained did not show any lower pH (e.g. pH 6) where the decomposition was not
solvolysis in the system we studied. affected by pressure. The optimum conditions for esters
The same effect also appears in the case of carboxylic are 800 MPa at pH 8 and for acids are either 400 or
acids and alcohol solutions. Carboxylic acids and 800 MPa at pH 8.
alcohols should react to produce esters when pressure Hydrogen bonding and compound volatility are also
is applied due to the oncoming decrease in volume. affected by pH conditions. Hydrogen bonding is affected
Moreover, interactions between the acid and the solvent only in the case of carboxylic acids because esters do not
lead to a further increase in volume. The acid interacts form this type of bonds. In this case, when pH decreases,
with water and hydrates. This hydration leads to a the degree of acid dissociation also decreases. Thus,
further increase in volume. [ROOH] becomes the predominant form. [ROOH] has
On the other hand, hydrogen bonds are formed among the ability to form more hydrogen bonds than [ROO]
the acid, the alcohol and water. The formation of species. As a result, acidic conditions enhance hydrogen
hydrogen bonds can lead to a small volume contraction. bond formation.
Consequently, when pressure is applied, a volume Ester and carboxylic acid volatility are both affected by
decrease can occur due to hydrogen bond formation pH. In the case of esters, the effect is rather complicated.
and not necessarily due to ester formation. As pH conditions become more acidic, hydrolysis is
In any case, the role of pH is very important in ester enhanced, ROOR0 concentration is decreased while
formation and hydrolysis and thus the selection of the ROOH concentration is increased. But ROOH species
experimental pH conditions is of great importance. In are relatively more volatile than ROOR0 species. So,
the case of base-induced hydrolysis, a mineral acid, for even though ester volatility is not decreased, esters are
instance HCl, is needed in the last step to protonate the hydrolysed into more volatile acids. A low-pressure
carboxylate ion and release the carboxylic acid. In the treatment in basic conditions is thus required to ensure
case of acid-catalysed hydrolysis, a protonation is minimum damage.
required to activate the carbonyl group during the first On the other hand, pH conditions affect carboxylic acids
step. In both cases, low pH values favour hydrolysis as in the same way as in hydrogen bond formation. In
the degree of protonation increases. As a result, low pH acidic conditions ROOH, which is the predominant
values favour hydrolysis. species, is more volatile than ROO species. So, as pH
In general, it seems that pH has the same effect on both decreases, acid volatility is increased.
formation and hydrolysis reactions. In low pH values Here, the reported decomposition (i.e. loss) could be
both reactions are favoured, while as pH increases the possibly attributed to the high volatility of the
rate of both reactions is decreased, expecting higher compounds used and the inherent experimental error.
rates of reaction in more acidic conditions. In all cases, both HHP treatments of 400 and 800 MPa
The obtained results indicate a dependency of decom- had no effect on the overall reaction. The degree of
position on pH. To be more specific, the degree of decomposition varied with pressure. Ester decomposi-
decomposition is decreased as pH increases. This is due tion minimised during 400 MPa HHP treatment, while
to the fact that in high pH values the rate of both acid decomposition appeared to be independent of the
reactions, hydrolysis and formation, is reduced. As a pressure applied.
result both ester and acid solutions were more stable in The results reported here could be explained on the basis
basic conditions. In the case of esters, the optimum that high pressure was applied for a short period of time
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(15 min) and thus it can be considered as a mild KIMURA, K., IDA, M., YOSIDA, Y., OHKI, K., FUKUMOTO, T.
treatment. The selection of the processing time was AND SAKUI, N. Comparison of keeping quality between
based on the previous work on high-pressure treatment. pressure-processed jam and heat processed jamFchanges in
flavor components, hue and nutrients during storage.
High pressure was applied for a period of 10–30 min to Bioscience Biotechnology and Biochemistry, 58, 1386–1391
strawberry jam (Kimura et al., 1994), 15 min to guava (1994)
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processing time in our work was selected as an average McMurry, J. Organic Chemistry, 4th Edn. New York: Brooks–
of the above. Cole Publishing Company, pp. 250–295 (1994)
Comparing the obtained results with that of the MERTENS, M. Hydrostatic pressure treatment of food:
previous research in this area, 400–600 MPa pressure equipment and processing. In: GOULD, G. W. (Ed), New
Methods of Food Preservation. London: Blackie Academic &
treatments seem to have no or minor effects. A 400 MPa Professional, pp. 136–150 (1995)
treatment on strawberry jam preserved the fresh flavour OHSHIMA, T., USHIO, H. AND KOIZUMI, C. High pressure
and natural colour (Kimura et al., 1994), 600 MPa processing of fish and fish products. Trends in Food Science
sterilised guava juice (Yen & Lin, 1999), 500 MPa and Technology, 4, 370–374 (1993)
preserved the original flavour and taste of fish (Ohshima SOHN, K. H. AND LEE, H. J. Effects of high-pressure treatment
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composition of aromatic compounds in peach (Sumitani Changes in composition of volatile compounds in high
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TAUSCHER, B. Pasteurisation of food by hydrostatic high
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highest flavour stability was observed when samples Cleavage of esters under nearly neutral conditions at high
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appreciated. ZABETAKIS, I., KOULENTIANOS, A., ORRUÑO, E. AND BOYES, I.
The effect of high pressure on strawberry flavour
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