Journal of Food Engineering 65 (2004) 165–173

www.elsevier.com/locate/jfoodeng

Isothermal calorimetry approach to evaluate tissue damagein carrot slices upon thermal processing

Federico G�omez a,*, Romeo T. Toledo b, Lars Wads€o c, Vassilis Gekas a,d,Ingegerd Sj€oholm a

a Department of Food Engineering, Lund University, P.O. Box 124, SE-221 00 Lund, Swedenb Department of Food Science and Technology, The University of Georgia, Athens, GA 30602, USA

c Department of Building Materials, Lund University, P.O. Box 118, SE-221 00 Lund, Swedend Department of Environmental Engineering, Technical University of Crete, Chania, Greece

Received 24 July 2003; accepted 5 January 2004

Abstract

The purpose of this investigation was to test calorimetry as a means of quantifying cell damage occurring at various stages of

mild blanching treatments. Calorimetric measurements were carried out on cores taken from carrot slices blanched at 100 �C for 5–

45 s. The calorimetric data were corrected for the influence of wounding stress due to slicing and coring. The validity of calorimetry

to measure cell damage was assessed by simulating inactivation of cell metabolism based on bibliographic data on thermal inac-

tivation kinetics of plasma membrane Hþ-ATPase, used as a marker of overall cell membrane damage. Calorimetric data paralleled

the simulated decrease of actively metabolizing cells only in the first 20 s of blanching but diverged at higher blanching times. Our

results suggest that inactivation of enzymes responsible for quality-related changes during frozen storage (e.g. blanching at 100 �Cfor 20 s) may be accompanied by cell damage in about 70% of the tissue.

� 2004 Elsevier Ltd. All rights reserved.

Keywords: Calorimetry; Mild blanching; Cell damage; Hþ-ATPase

1. Introduction

In the frozen vegetable industry, blanching is used to

inactivate enzymes that cause off-flavour, changes in

colour and other chemical reactions during frozen

storage. Several blanching times and temperatures have

been reported for blanching carrot slices of various

thickness. These include treatments in boiling water (100

�C), 3–4 min (3–5 mm-thick slices) (Lee, Bourne, & VanBuren, 1979; Pr�estamo, Fuster, & Risue~no, 1998; Rah-

man, Henning, & Westcott, 1971) and 90 �C, 4 min

(Kidmose & Martens, 1999). These treatments were di-

rected towards the inactivation of peroxidase, however,

the use of this enzyme as an indicator of the blanching

process may result in undesirable quality changes (Kim

et al., 2001). The use of other enzymes as indicators of

the blanching process has also been suggested as a meanto minimise the heat treatment. Roy, Taylor, and Kra-

*Corresponding author. Tel.: +46-46-222-98-22; fax: +46-46-222-46

22.

E-mail address: federico.gomez@livstek.lth.se (F. G�omez).

0260-8774/$ - see front matter � 2004 Elsevier Ltd. All rights reserved.

doi:10.1016/j.jfoodeng.2004.01.010

mer (2001) used lipoxygenase as a marker, obtaininggood retention of quality attributes after blanching 1-cm

carrot cubes at 100 �C for 0.58 min. These blanching

procedures, following a classical approach, are all based

on the average effect of the heat treatment in the sample.

Mathematical modelling for blanching optimization

has been directed towards a better understanding of

processes taking place during the heat treatment such as

turgor loss (De Belie, Herppich, & Baerdemaeker, 2000)and starch gelatinization (Verlinden & De Baerdemae-

ker, 1997). These models were based on a detailed

description of the temperature distribution in the sam-

ple, from the surface towards the centre. However, as

the thermal properties of the samples are usually taken

from the literature at a low, constant temperature, the

models failed to consider changes in these properties

during the heating process. Chang and Toledo (1990)reported a significant elevation of effective thermal dif-

fusivity as a result of internal convective heat transfer

due to penetration of heating fluid into the sample and

possible structural changes. Ignoring changes of thermal

conductivity with temperature would cause errors in the

Nomenclature

Subindex i slice section

Superscript k time interval

a thermal difussivity (m2 s�1)

b calibration factor of the line heat source

probe used for determination of thermalconductivity

� calibration coefficient of the calorimeter

(mWmV�1)

q density (kgm�3)

ln term in Eq. (6)

C fraction of thermal power contributed by

normal cell metabolic activity

Cd fraction of actively metabolizing cells (un-damaged and partially damaged) ¼ remained

activity of plasma membrane Hþ-ATPase

(1/s)

Cp specific heat (J kg�1 K�1)

I thermal conductivity probe heater input cur-

rent (A)

k thermal conductivity (Wm�1 K�1)

kd first order kinetics constant of inactivation ofplasma membrane Hþ-ATPase (1/s)

k0 kinetic constant of Arrhenius law ({}/s)

Ea activation energy (KJmol�1)

M mass of the sample (g)

n numerical index

P thermal power (mWg�1)

r variable distance from the centre axis of the

slab (m)R gas constant (8.314 J gmol�1 K�1)

Rk resistance/length of Constantan heating wire

for thermal conductivity measurement (X/m)

RS half-thickness of slab (m)

S slope of the linear portion of a plot temper-

ature vs. ln time

t time (s)

T temperature (�C)TM temperature of the blanching water (�C)T0 initial temperature (�C)VS voltage recorded from the sample in the cal-

orimeter (mV)

VBL voltage recorded from the base line in the

calorimeter (mV)

X thickness of slice (mm)

XW fraction of water contentW fraction of the total thermal power contri-

buted by the reaction to wounding stress

166 F. G�omez et al. / Journal of Food Engineering 65 (2004) 165–173

temperature profile calculations (Gratzek & Toledo,

1993) and in the proposed blanching models.

Starting from the surface of the heated sample,

blanching will lead to cell membrane disruption and

protein denaturation (Hansen, Afzal, Breidenbach, &

Criddle, 1994). Other processes such as turgor loss and

starch gelatinization will be a direct consequence of

membrane damage (Greve et al., 1994), influencingquality attributes such as firmness and crispness (De

Belie et al., 2000). Possibilities of mild blanching con-

ditions have been explored for preservation of tissue

integrity and quality improvement (Pala, 1983; Pizzo-

caro, Senesi, Querro, & Gasparoli, 1995). Mild blanch-

ing treatments directed towards avoidance of oxidative

phenomena in carrot lipids during frozen storage were

applied at temperatures as high as 100 �C for a fewseconds, a process designed to inactivate enzymes near

the surface of the carrot pieces. Samples kept in boiling

water for 20–30 s followed by immediate cooling in tap

water, appeared to exhibit fresh-like quality after frozen

storage (Pizzocaro et al., 1995), therefore, the process

may be potentially beneficial in minimising cellular

damage so that sensorial quality in the stored frozen

carrots is maximised. However, there are to ourknowledge no previous reports on how to measure the

extent of tissue damage during these mild blanching

treatments that can be used to set the optimal range of

the treatments.

Inactivation of cell metabolism, and hence, cell

damage due to high temperature injury has been as-

sessed using calorimetry (Hansen et al., 1994; Rank,

Breidenbach, Fontana, Hansen, & Criddle, 1991).

Calorimetric measures of the rate of heat production are

proportional to metabolic rates and provide a directindication of integrated metabolic responses such as

respiration and reaction to wounding stress (Criddle,

Breindenbach, & Hansen, 1991; Iversen, Wilhelmsen, &

Criddle, 1989). Calorimetry may be potentially useful to

quantify cell damage that occurs during processing

operations such as blanching. Increasing intensity of the

heat treatment would progressively reduce the number

of viable cells in the tissue and thus, result in a corre-sponding reduction in the rate of metabolic processes.

The purpose of this investigation has been to test

calorimetry as a means of quantifying the decrease of

cell viability in carrot tissue during processing and

hence, to evaluate tissue damage occurring at various

stages of the blanching treatment. A simulation was

carried out to calculate the extent of cell membrane

damage at different times of blanching at 100 �C basedon thermal inactivation kinetics of plasma membrane

Hþ-ATPase as a marker of overall membrane damage.

F. G�omez et al. / Journal of Food Engineering 65 (2004) 165–173 167

Plasma membrane Hþ-ATPase actively pumps protonsout of the cell creating pH and electrical potential dif-

ferences across the membrane. Many physiological

processes are driven by this electrochemical proton

gradient such as the transport of solutes into and out of

the cell. The enzyme also has an important role in reg-

ulating the cytoplasmic pH, and the pH gradient

developed over the membrane may also affect pH-sen-

sitive enzymes and plasma membrane channel proteins(Michelet & Bountry, 1995). Hþ-ATPase enzyme activ-

ity reflects both the physical and the physiological state

of the cell membranes (Gonz�alez-Mart�ınez, 2003; Stan-ley, 1991). Thus, by using thermal inactivation kinetics

of Hþ-ATPase, we attempted to have a marker of

overall membrane damage due to thermal process, as

membranes will lose their capacity to control the active

transport of ions and solutes into and out the cytoplasm(Gonz�alez-Mart�ınez, 2003).

2. Materials and methods

2.1. Raw material, handling and storage

Carrots (Daucus carota L. cv. Nerac), grown in the

south of Sweden, were used. The carrots, 400 kg, har-

vested in the last week of October 2001, were washed,

hydrocooled to about 3 �C and stored in one largepolyethylene plastic bag at 0 �C and approx. 100% rel-

ative humidity, as routinely practiced by the producer.

Medium-sized carrots (19.5 ± 1.0 cm in length, 3.0 ± 0.2

cm diameter) free from defects, were selected at the farm

storage facility and transported to our laboratory in

plastic bags kept on ice in insulated boxes. The carrots

were then taken out of the bags and placed in a closed,

refrigerated chamber at )0.2 �C and 100% relativehumidity until used.

2.2. Preparation of samples

Carrots were manually peeled, the top and tip re-

moved and the remainder was cut into slices. Slices 3.0–

3.2 cm diameter and 7 mm thick were selected and used

for the blanching experiments. After blanching, four tap

root cores 7 mm long and 11 mm in diameter were ex-cised longitudinally from the phloem parenchyma of

each slice with a cork borer. The cores were used for

isothermal calorimetry measurements and evaluation of

damaged cells with Trypan blue staining.

2.3. Blanching

Carrot slices was immersed in boiling water (100 �C)for a defined time and rapidly cooled in ice water to

minimise over-processing. Non-heated slices of the same

dimensions were used as control.

2.4. Isothermal calorimetry measurements on the

blanched samples

Isothermal calorimetry was performed with a proto-

type of the TAM Air isothermal calorimeter (Thermo-

metric AB, J€arf€alla, Sweden). This isothermal heat

conduction instrument contains eight twin calorimeters.

The instrument was placed inside an insulated holder that

was placed in a cooled incubator. This arrangement gaveenough temperature stability for the present measure-

ments. The samples were placed in 20 ml glass ampoules

sealed with teflon-coated rubber seals and aluminium

caps. Each calorimeter had its own reference which

consisted of a sealed 20 ml glass ampoule containing 4 ml

water. The eight calorimeters permitted eight simulta-

neous measurements of the rate of heat production.

Samples for calorimetry were four cores obtainedfrom the phloem parenchyma of raw and blanched

carrot slices. To perform the isothermal calorimetry

measurement, each sample was placed in a numbered

ampoule that was immediately weighed and sealed. In

this way samples consisting of a raw sample (control)

and seven other heat treated slices, were distributed in

eight ampoules. The sealed ampoules were immediately

placed in the calorimeters.The isothermal measurements were performed at 20

�C for a period of about 24 h. Baselines (BLs) were re-

corded before or after each measurement. The primary

output from the heat flow sensors in the calorimeters (a

voltage) was recorded by computer from the digitized

output of the calorimeters. The corresponding thermal

powers (heat production rates) were calculated by the

following equation:

P ¼ e�VS � VBL

Mð1Þ

were P (mWg�1) is the specific thermal power of the

carrot sample, e is the calibration coefficient of the cal-

orimeter (mWmV�1), VS is the voltage (mV) recorded

from the sample, VBL is the voltage (mV) recorded from

the base line and M is the mass (g) of the sample. Thecalibration coefficients were calculated from electrical

calibrations made at 20 �C.

2.5. Isothermal calorimetry measurements on the

blanched samples: influence of wounding on thermal power

When tissues are wounded, the cells at the site of such

a stress strengthen their walls by the secretion of addi-

tional structural components such as hydroxyproline-

rich glycoproteins, lignin or suberin (Satoh, Sturm,

Fujii, & Crispeels, 1992). To evaluate the metabolic heat

produced by these stressed cells near the cut surface andthus, estimate the contribution of wounding stress to the

total thermal power measured in the blanched samples,

a separate calorimetric experiment was carried out. Four

168 F. G�omez et al. / Journal of Food Engineering 65 (2004) 165–173

carrot root cores were submerged in boiling water forabout 2 s, a treatment that will damage only the cells

near the cut surface of the samples, and immediately

cooled in ice water. To perform the isothermal mea-

surement, the cores were placed in a numbered ampoule

which was immediately weighed and sealed. Four cores

of the same dimensions, taken from another slice of the

same carrot were kept raw as control and placed in a

second ampoule. The heat production rate from eachampoule was measured in a TAM Air instrument as

described previously for the blanched samples. The

difference in the thermal power between both samples,

as calculated with Eq. (1), will be a direct estimation of

the heat contributed by the cells near the cut surface.

The same experiment was repeated for each of the

blanching treatments. Four cores were obtained from

each of the blanched slices and placed in a calorimeterampoule. Four cores of the same dimensions, taken

from another slice, blanched for the same period of time,

were heated for about two additional seconds to damage

the cells from the surfaces that were not in direct contact

with the boiling water when the entire slice was blan-

ched, and placed in a second ampoule.

2.6. Evaluation of damaged cells using trypan blue

staining solution

Carrot root cores from the phloem parenchyma of

each slice used in the blanching experiment were treated

with Trypan blue staining solution (0.4%, Sigma-Al-

drich, USA). Trypan blue is a membrane-impermeabledye, which is not taken up by viable cells which have an

intact plasma membrane. Thus, damaged cells (non-

viable) stain and living cells (viable) do not (Danielson

et al., 2002). The cores were steeped 5 min in the staining

solution, washed with distilled water to remove excess of

dye and examined under a stereomicroscope (Olympus

SZ-CTV, Japan).

2.7. Determination of thermal and physical properties

The thermal conductivity, k (Wm�1 K�1), density, q(kgm�3) and moisture content were experimentally

determined and used to calculate heat penetration

through the slices during blanching treatments.The thermal conductivity was measured using a line

heat source probe as described by Gratzek and Toledo

(1993) with some modifications on the power supply and

measurement circuit. The probe was calibrated by

immersion in glycerine (k ¼ 0:2873 Wm�1 K�1) Carrots

with more or less uniform cylindrical shape were se-

lected, peeled and top and tip separated. The thermal

conductivity probe (with Teflon insulated Constantanheater wire and Teflon insulated Copper–Constantan

thermocouple wire enclosed in a 6.9 cm long, 1.27 mm

diameter stainless steel active probe section) was in-

serted along the axis of the 7.3 cm-long, cylindricalcarrot samples. The carrots, placed in a plastic mesh,

were carefully immersed in a thermostatic bath set at 20

�C. The temperature of both the surrounding fluid and

the sample were monitored until they were identical.

When the sample and surrounding water were at the

same temperature, the measurement was started. The

measurements were done by admitting a pre-set constant

current into the probe heating wire. The current levelwas determined by measurement of the voltage drop

across a high precision (0.05%) 1.0 ohm shunt resistor

with a digital voltimeter. A current level of 0.140 A was

used for all tests. The temperature signal was measured

at a sampling frequency of 200 Hz with a time averaging

each 0.15 s. Tests were usually run for 30 s. Time and

temperature information was recorded in a data acqui-

sition system (Fluke, USA).Thermal conductivity, k, was calculated from the

following (Gratzek & Toledo, 1993):

k ¼ b2I2Rk

4pSð2Þ

where IðAÞ is the probe heater input current, Rk (X/m) isthe resistance/length of Constantan heating wire, and bthe calibration factor determined during the calibration

of the line heat source probe with glycerin. The

numerator is multiplied by 2 because the wire doubled

back in the probe. S is the slope of the linear portion of a

plot DT vs. ln time. DT is the temperature rise from the

equilibrium temperature as recorded by the computer

from the output of the probe thermocouple.The change of thermal conductivity with increasing

temperature was measured. When the temperature of

the surrounding water was increased, the temperature of

the sample was monitored until equilibrium was

reached, at which time a new thermal conductivity

measurement was initiated. After a pause to re-establish

equilibrium, another measurement was made at the

same temperature. After completing measurements at aparticular temperature, the water was heated and the

procedure repeated at the next temperature level.

The density of carrot pieces was measured at 20 �Cwith an ultrapycnometer 1000 (Quantachrome, USA)

using helium as the fluid. Sample moisture was deter-

mined after drying the sample (ca 3 g fresh weight) for

18 h in a vacuum-oven (VWR Scientific Products, USA)

at 70 �C and 380 mm Hg of absolute pressure.

3. Results and discussion

3.1. The thermal power decreases as the intensity of the

blanching process increases

An example of the results from the calorimetric

measurements of raw and blanched carrots is shown in

F. G�omez et al. / Journal of Food Engineering 65 (2004) 165–173 169

Fig. 1a. Each time an ampoule was charged into acalorimeter there was an initial disturbance lasting for at

least 30 min. After that, the true voltage (mV) from the

samples was measured. In the unblanched sample (0 s),

the voltage increased and then reached a short plateau

after about 8 h, followed by a decrease that could be

attributed to the depletion of oxygen in the ampoule

(Criddle, Breidenbach, Lewis, Eatough, & Hansen,

1988). In the 5 and 10 s-blanched samples, the voltageincreased during the first 8 h of the measurement al-

though it did not increase as much as that of the raw

sample. The following plateau was fairly constant until

the experiment was interrupted. In the 20, 30 and 45 s-

blanched samples the voltage did not increase at the

beginning of the measurements and was fairly constant

during the time course of the experiment with a slight

tendency to increase in the last hours. The curve corre-sponding to the 45 s-blanched sample was fairly con-

stant and very close to that of the baseline. Interestingly,

there were remarkable differences in the voltage values

Fig. 1. (a) Calorimeter output of blanched carrot samples at 100 �C for

different time intervals. Blanching time is indicated next to each curve.

The experiment was interrupted after 24 h and the baseline (BL) was

recorded. (b) Relative thermal power (heat production rates) of carrots

with increasing blanching time. The thermal power, P , calculated with

Eq. (1) was normalized by dividing values with those of the raw carrot

(control). Means of three measurements are shown.

from the different samples, decreasing with increasingblanching time. The experiment was interrupted after 24

h by taking the samples out from the calorimeter. This

interruption caused a disturbance lasting for at least 30

min before the BL started to be recorded.

The constant voltage plateau of the curves was used

to calculate thermal power according to Eq. (1). Fig. 1b

describes the decrease in the thermal power of carrots

blanched at 100 �C for different time intervals.The thermal power of the carrot samples decreased

sharply during the first 20 s of blanching and then re-

ceded slowly with prolonged heating to reach a very low

value, 1.6% after 45 s. Since the thermal power is a

parameter related with metabolic activity of the cells in

the tissue (Criddle et al., 1991), our results suggests that

the amount of living cells by that time is very low or

null. In support of this idea, Fig. 2 shows that the carrottissue blanched 45 s exhibited complete staining with the

Trypan blue, indicating that viable cells are no longer

present. This result is in agreement with Greve et al.

(1994) who could not identify any living cells in 1 cm-

thick carrot slices blanched at 100 �C for 60 s using a

flourescein diacetate dye.

3.2. Estimation of the reaction to wounding stress

We could consider the thermal power values (P in

Fig. 1b) to be the result of the sum of the energy released

by two main physiological processes, that from the

‘‘normal’’ cell metabolic activity ðCÞ and that originat-

ing from wounding stress produced by the cells near the

cut surface of the cores ðW Þ:P ¼ W þ C ð3Þ

where W is the result of the experimental estimation ofthe heat contributed by the cells near the cut surface.

The contribution of wounding stress to the thermal

power is summarised in Table 1. The results show that

carrot metabolism is highly influenced by the wounding

reaction. In the raw carrot sample, the contribution of

the reaction to wounding stress to the total thermal

power was 52% and, surprisingly, in the 30 s-blanched

samples this contribution was 41%.

3.3. Simulation of cell damage during blanching treat-

ments

3.3.1. Kinetic parameters

Cell membrane damage upon heating has been re-

cently investigated by Gonz�alez-Mart�ınez (2003). The

authors used vesicles isolated from potato cell mem-

brane to test inactivation of plasma membrane Hþ-ATPase activity as a marker for cell membrane damage.

Eppendorf tubes containing isolated cell membranes in

suspension were heated at selected temperatures and

Table 1

Fractions of ‘‘normal’’ cellular metabolism, C and wounding stress, Wof the total thermal power (heat production rates), P (plotted in Fig.

1b), measured with isothermal calorimetry in cores taken from carrot

slices blanched at 100 �C for various time intervals

Blanch-

ing time

(s)

P (mW) C1 W a P �C(mW)

0 0.3908 0.484 0.516 0.189

5 0.2462 0.498 0.502 0.132

10 0.1911 0.552 0.448 0.105

20 0.0824 0.559 0.441 0.046

30 0.0469 0.595 0.405 0.028

45 0.007 1.000 – 0.007

a The values are the mean of duplicate determinations; variations

within duplicates were <5%.

Fig. 2. Front and side of cores obtained from carrot slices blanched at 100 �C for different times and stained with Trypan blue, as indicated in the

Materials and methods section. Damaged cells were stained (dark areas) while intact cells did not absorb the stain.

170 F. G�omez et al. / Journal of Food Engineering 65 (2004) 165–173

times and the Hþ-ATPase activity was measured after

the heat treatment to determine the inactivation kinetics

of the enzyme. The authors determined Hþ-ATPase

activity at 20 �C according to Palmgren and Sommarin(1989). In this method, ATP hydrolysis is coupled

enzymatically to the oxidation of NADH and the rate of

hydrolysis is measured as the decrease in absorbance of

NADH at 340 nm. The activity was determined after the

addition of 0.05% (w/v) polyoxyethylene-20-cetylether

(Brij 58) that produced 100% inside-out vesicles suitable

for the activity measurements.

The kinetics of inactivation of this enzyme showed tobe first order:

dCd

dt¼ �kdCd ð4Þ

The temperature dependence of the kinetics constant kdcould be expressed by two linear Arrhenius plots:

kd ¼ koe�EaRT ð5Þ

The first was valid in the range between 45 and 53 �Cand the second was valid in the range between 60 and 75

�C. The authors calculated 55 �C as a transition tem-

perature represented by the intercept of the two linear

plots. The kinetic parameters for inactivation of Hþ-

ATPase are summarised in Table 2. We here assume the

inactivation of Hþ-ATPase, an indicator of cell mem-brane damage, to be proportional to the decrease in cell

metabolic activity.

3.3.2. Calculation of temperature profile

The temperature distribution in the carrot slice dur-

ing the blanching treatments was calculated as follows.

The carrot slice, considered as an infinite slab, was

divided horizontally, from the surface to the centre, into

7 sections of 0.5 mm each. The change of temperature, T(�C) with time, tðsÞ, in the centre of each of the slabsections during blanching, was calculated with the fol-

lowing equation (Singh, 1992):

T � TMT0 � TM

¼X1n¼1

2

lnð�1Þnþ1

coslnrRS

exp

�� l2

n

atR2S

�ð6Þ

where RS (m) is the half-thickness of a slab and r (m) isthe variable distance from the centre axis, TM is the

temperature of the blanching water (100 �C), T0 is the

initial temperature (20 �C), ln is defined by

Table 2

Parameters of the first-order kinetics for inactivation of Hþ-ATPase in

plasma membranes isolated from potato tubers (adapted from

Gonz�alez-Mart�ınez (2003), Fig. 2)

Temperature (�C) kd (1/s)

45 0.070

49 0.110

53 0.244

60 0.372

38 0.372

75 0.507

F. G�omez et al. / Journal of Food Engineering 65 (2004) 165–173 171

ln ¼ ð2n� 1Þ p2

ð7Þ

a, the thermal difussivity (m2 s�1), is calculated by

a ¼ kqCp

ð8Þ

where the increase of the thermal conductivity, k, withtemperature (Fig. 3) was measured as

k ¼ 0:5464þ 0:0012T � 2:0� 10�6T 2 ðr2 ¼ 0:9838Þð9Þ

The measured density, q ¼ 1154 kgm�3 and the specific

heat, Cp ¼ 3736 J kg�1 K�1 as calculated with the mois-

ture content of the carrots according to Siebel (1918):

Cp ¼ 0:837þ 3348 Xw ð10Þ

The summatory term in Eq. (6) was repeated three

times.

93.7˚ C81.2˚ C69.4˚ C59.1˚ C50.8˚ C45.1˚ C42.1˚ C

Xi = 0.5 mm

˚

˚˚˚˚˚

100˚C

3.3.3. Decrease of cell metabolic activity during blanching

The effect of the temperature distribution in the car-

rot sample during blanching treatments on the decrease

of cell metabolic activity was investigated as follows.

The fraction of actively metabolizing cells (undam-aged and partially damaged) Cdi at the centre of each of

the slice sections i as a function of time (superscript k)was calculated by solving Eq. (4):

0.540

0.560

0.580

0.600

0.620

0.640

10 30 50 70 90

k (w

/m°K

)

Temperature (°C)

Fig. 3. Carrot thermal conductivity as a function of temperature.

Ckþ1di ¼ Ck

di � kT ki

d CkdiDt ð11Þ

where kd is a function of the temperature at the centre of

each element i at time k, according to Eq. (5). Fig. 4a

and b shows an example of the result of the calculation,showing both the values of temperature and the fraction

of actively metabolizing cells, Cdi in the centre of each of

the slice sections of thickness X after 20 s of blanching at

100 �C.The fraction of actively metabolizing cells, Cd , in the

whole slab is obtained by averaging all Cdi at each time

step:

Cd ¼2P

XiCdi

Xð12Þ

3.4. Fitting the calorimetric data to the simulated decrease

of cell metabolic activity

The values of thermal power P , plotted in Fig. 1b and

reported in Table 1, were multiplied by the fraction ofthe ‘‘normal’’ cell metabolic activity, C (Table 1), to

obtain the value of thermal power that excludes the heat

produced by the reaction to wounding stress of the cells

near the cut surface. The resulting values (P �C in Table

1) are plotted in Fig. 5, expressed as relative to the one

of the control. These values are fitted in the same figure

to the fraction of actively metabolizing cells, Cd .

Fig. 5 shows that the decrease of cell metabolicactivity simulated using the inactivation kinetics of Hþ-

ATPase parallels the calorimetric data adequately for

the first 20 s of blanching. The error bars enclose the

Centre

Surface

0

00.0075

0.0956

0.53960.8144

0.9012

Centre

Xi = 0.5 mm

(b)

(a)

Fig. 4. (a) Schematic representation of the calculated temperature

distribution in a carrot slice after 20 s of blanching in water at 100 �C.(b) Schematic representation of the distribution of the fraction of ac-

tively metabolizing cells, Cdi, in a carrot slice after 20 s of blanching in

water at 100 �C (calculated from temperature distribution and thermal

inactivation kinetics of Hþ-ATPase).

0

20

40

60

80

100

0 10 20 30 40 50

Blanching time (s)

P * C

(% o

f con

trol

)

0

10

20

30

40

50

60

70

Cen

tre

tem

pera

ture

(°C

) (

)

Cd

(

)

0

1

Fig. 5. Comparison of calorimetrically measured metabolic heat pro-

duction of blanched carrot slices, P �C in Table 1 (filled circles), and

fraction of actively metabolizing cells, Cd , simulated from carrot

temperature profile and literature data on Hþ-ATPase inactivation

kinetics (solid line). Data on Hþ-ATPase inactivation kinetics, re-

ported in Table 2, was obtained from Gonz�alez-Mart�ınez (2003). The

temperature at the centre of the slices is depicted by the dashed line.

172 F. G�omez et al. / Journal of Food Engineering 65 (2004) 165–173

simulation line during that period of time except for the

treatment for 5 s. At higher blanching times the simu-

lated Hþ-ATPase inactivation and calorimetric data

were highly divergent.The optimal range of blanching should be set based

on the inactivation of enzymes responsible for the

development of off-flavours for example in the frozen

carrots during storage. Activity of catalase was used as a

marker for adequate blanching of carrots by Pizzocaro,

Aggujaro, and Bertolo (1993). These authors reported a

loss of about 56% of initial catalase activity after

blanching 0.6 cm-thick carrot slices at 100 �C for 10 s,90% after 20 s and 94% after 30 s. Only the samples

blanched for 10 s developed off-flavours during frozen

storage at )20 �C. Our results suggest that inactivation

of enzymes responsible for the development of off-fla-

vours during frozen storage (e.g. blanching at 100 �C for

20 s) may be accompanied by inactivation of cell

metabolism, and hence, cell damage in about 70% of the

tissue.

4. Concluding remarks

Our simulation described well the isothermal calo-

rimetry measurements during the first 20 s of blanching,

where a sharp decrease in the thermal power occurred,

suggesting the potential of calorimetry as a means of

quantifying cell damage. The use of markers for study-

ing the inactivation of cell metabolism during heat

treatment is an issue worthy of future research. We here

have used bibliographic data on inactivation of Hþ-ATPase, measured as the capacity of the enzyme to

hydrolyze ATP. As this capacity is impaired, inactiva-

tion of Hþ-ATPase could be considered as a good

marker for membrane damage. However, this mea-surement does not give any indication of the capacity of

the enzyme to actively pump protons out of the cell.

Gonz�alez-Mart�ınez (2003) suggested that the capacity

for proton pumping begins to be affected at lower tem-

peratures than the capacity to hydrolyze ATP. Loss of

proton pumping capacity will be a marker of cell

membrane dysfunction and the thermal inactivation

kinetics of this event could also be considered as a goodmarker. Activity of mitochondria, the ‘‘energy genera-

tor’’ of the cell might also be a good marker of loss of

metabolic activity.

Actual data on thermal inactivation of one or more of

these markers in carrot cells may give closer correlation

with calorimetric data. We know very little about dif-

ferent factors that may contribute to the thermal power

values after 30 s of blanching when most of the cells arealready damaged.

Future work correlating inactivation of metabolic

activity with quality parameters such as texture and loss

of nutrients will be important for the optimization of

blanching treatments aimed to preserve tissue integrity

and improve the quality of the final product.

Acknowledgements

The authors would like to thank Prof. Marianne

Sommarin, Department of Plant Biochemistry, Lund

University, Sweden for enlightening discussions. We

also wish to acknowledge our thanks to Marianne’sFarm for supplying the carrots used in this study. This

study was supported by grants from: The Swedish Na-

tional Board for Industrial and Technical Development

(NUTEK); The Centre for Nutrition Research (LCL);

The Swedish Research Council (MS); the Foundation

‘‘Marknads Tekniskt Centrum’’; The Scanian Dairies

(The Economic Association of the Dairy Cooperation),

The Scanian Farmers’ Cooperation and The RoyalPhysiographic Society in Lund, Sweden.

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