Journal of Food Engineering 148 (2015) 1323

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Journal of Food Engineering

journal homepage: www.elsevier.com/locate/jfoodeng

Kinetic study of quality indices and shelf life modelling of frozen spinach
under dynamic conditions of the cold chain
E. Dermesonluoglu a, G. Katsaros a, M. Tsevdou a, M. Giannakourou b, P. Taoukis a,
a
b

Laboratory of Food Chemistry and Technology, School of Chemical Engineering, National Technical University of Athens, Iroon Polytechniou 9, Zografou 15780, Athens, Greece
Department of Food Technology, Faculty of Food Technology and Nutrition, Technological Educational Institute of Athens, Ag. Spyridonos, Aigaleo 12243, Athens, Greece

a r t i c l e

i n f o

Article history:
Received 24 March 2014
Received in revised form 7 July 2014
Accepted 13 July 2014
Available online 29 July 2014
Keywords:
Spinach
Non-isothermal frozen storage
Vitamin C
Chlorophyll
Texture
Sensory
Shelf life

a b s t r a c t
A systematic kinetic study of quality loss of frozen spinach was conducted. Vitamin C, chlorophyll (a, b
and total), colour, texture and sensory attributes were measured in frozen leafy spinach. Kinetic models
of the selected quality indices were developed to determine the frozen spinach quality loss and calculate
product remaining shelf life. Validated kinetic models of quality loss are applied in the evaluation, control
and management of frozen spinach in the cold chain. Storage experiments were conducted at isothermal
(5, 8, 12, 18 C) and non-isothermal temperature conditions (Teff = 6.9 C) to validate proposed
shelf life models. The temperature dependence of quality loss was adequately modelled by Arrhenius
type equation and activation energy values of 132, 72, 43 and 61 kJ/mol for vitamin C loss, total chlorophyll retention, texture deterioration and overall sensory scoring were calculated respectively. The shelf
life at the reference temperature of 18 C ranged from 400 to 500 days based on the different quality
indices. The effects of temperature deviations were quantitatively evaluated and a realistic assessment
of the frozen spinach remaining shelf life in the actual cold chain was conducted.
2014 Elsevier Ltd. All rights reserved.

1. Introduction
One of the widely consumed vegetables considered to have a
high nutritional value is spinach (Spinaciaoleracea L.). It contains
considerable amounts of vitamins, especially C, E, b-carotens, folic
acid, mineral components and dietary bres (Lisiewska et al.,
2009). Among the vegetables it is also characterized by its high
antioxidative capacity (Jaworska, 2005). Frozen spinach is preferred by most consumers due to its prolonged shelf life which
enables it to be available throughout the year. Its value as raw
material for freezing has been conrmed by numerous data in
the literature (Kmiecik and Lisiewska, 1999; Jaworska and
Kmiecik, 2000).
Inevitably, the freezing process and the frozen storage have
negative impacts on food quality. The main physical changes that
occur during frozen storage are moisture migration and ice recrystallization (Van Buggenhout et al., 2006). Both phenomena are
related to the stability of products frozen water, which affects
the vegetable texture and loss of nutrients and weight if thawing
drip loss takes place (Pukszta and Palich, 2007). Moreover, chemical changes in frozen vegetables cause off-avour development,

Corresponding author.
E-mail address: taoukis@chemeng.ntua.gr (P. Taoukis).
http://dx.doi.org/10.1016/j.jfoodeng.2014.07.007
0260-8774/ 2014 Elsevier Ltd. All rights reserved.

pigment and colour degradation (Lisiewska et al., 2009), oxidative

deterioration and vitamin loss (e.g. of vitamin C; Serpen et al.,
2007). Quality of frozen food is highly dependent on storage temperature (Taoukis et al., 1991). The knowledge of the exact inuence on storage temperature of frozen vegetables would help to
determine adequate conditions to preserve high quality and determine shelf life. The shelf life of frozen vegetables is normally established by sensory evaluation (IIR, 1986). Analytical measurements,
of colour and texture, and content of nutritional compounds, e.g.
vitamin C have been used in parallel to reinforce the sensory evaluation (Giannakourou and Taoukis, 2003; Reid et al., 2003; Martins
and Silva, 2002). However no uniformly applicable criteria have
been established and it is important to identify the shelf life determining factors for specic frozen vegetable.
Published work is available with regards quality degradation
and shelf life of frozen spinach. Vitamin C has been used as the
shelf life index being more susceptible to loss (L-ascorbic oxidation) during processing and storage (Favell, 1998; Giannakourou
and Taoukis, 2003). Crucial to the attractive appearance is the
green colour which depends on the amount of chlorophylls and
the ratio of chlorophyll a to chlorophyll b. The stability of chlorophylls in spinach subjected to different storage conditions has been
addressed only in few studies (Bunea et al., 2008). The colour
change has been expressed measuring the chlorophyll a and b,
and pheophytin a and b contents (Martins and Silva, 1998, 2002)

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as the major pathway of degradation of chlorophylls is by direct or

indirect pheophitisation to pheophytins. However, no consistent
relationship between chlorophyll losses and colour change has
been reported. Objective instrumental quantication of the colour
change based on measurement of CIELab values with the use of
chromatometer has been successfully applied for monitoring frozen vegetable quality loss (e.g. green peas, green beans)
(Giannakourou and Taoukis, 2002; Martins and Silva, 2002). Contribution of enzymatic activities (e.g. polygalacturonases, cellulases and hemicellulases) to textural changes in spinach has been
reported (Labib et al., 1997). Texture properties can be described
instrumentally with the use of texture analyzer measurements
and water holding capacity measurements (the change of weight
due to water absorbed or adsorbed) (Giannakourou and Taoukis,
2002; Martins and Silva, 2002). The enzymes responsible for loss
of desirable avour and aroma or formation of undesirable avour
are more difcult to pinpoint because of the complexity of avour.
The study of processing and storage effects on spinach avour,
aroma, colour and texture can be conducted by sensory evaluation
tests. No studies have been reported for the total quality loss of frozen spinach taking into account the nutritional content as well as
the sensory properties. In published work quality parameters
(e.g. vitamin C content) have been studied at constant low temperatures, indicative of frozen practice (e.g. at 20, 30 C), usually at
the beginning, in the middle and at the end of their commercial
shelf life, without assuming a full, systematic kinetic approach.
The temperature range in most studies does not cover the 3 to
10 C range which is very detrimental and can occur in the real
frozen chain (European Union survey EE1080/94/00069, 1995;
Giannakourou and Taoukis, 2002).
The objective of this work was to perform a thorough kinetic
study of different quality indices (vitamin C, chlorophyll (a, b and
total), colour, texture and overall acceptability) of frozen leafy
spinach, in a wide temperature range of frozen storage. Kinetic
models of the selected quality factors were developed to determine
the frozen spinach quality loss and calculate product remaining
shelf life. In order to validate these shelf life models, storage experiments were conducted at isothermal and non-isothermal conditions. The purpose of this validation, is to establish models
applicability in the real distribution chain and be able to reliably
estimate products remaining shelf life during their real lifecycle.
Using realistic case studies, the aim is to stress the importance of
knowing the real timetemperature history of the product and
underline the weakness of giving a unique, nominal shelf life
for all products. Furthermore, it will be clearly shown that the
choice of the quality parameter that signals products end of shelf
life is of major importance.

2. Materials and methods

2.1. Sample storage
Spinach leaf samples were supplied directly from the production line of a leading frozen vegetable product manufacturer. Samples were stored in their commercial packaging (laminate lm of
20 lm bio-oriented polypropylene(BOPP)-48 lm polyethylene(PE)) at temperature controlled cabinets (Sanyo MIR 153, Sanyo
Electric Co., Ora-Gun, Gunma, Japan) set at 5, 8, 12, 18 C
(isothermal temperature conditions) and at (non-isothermal) variable temperature conditions (12 h at 5 C, 12 h at 8 C and 12 h
at 10 C). The experiment under dynamic temperature conditions
was realized in order to validate the kinetic models developed
applying isothermal conditions. The specic dynamic prole was
chosen because it has an effective temperature close to 7 C,
which is well within the temperature range studied. The

temperature was constantly monitored by electronic, programmable, miniature dataloggers (COX TRACER, Belmont, NC) in the
incubators. Dataloggers were placed inside the incubators as well
as within one product package. Temperature standard deviations
at the isothermal conditions did not exceed throughout the experiment 0.2 C. Samples were also stored at 30 C as controls.
2.2. Vitamin C determination
Vitamin C (L-ascorbic acid) was determined using a high performance liquid chromatography method (HPLC) (Giannakourou and
Taoukis, 2003). All analyses were carried out in duplicate on vegetable tissue, homogenized, using a pestle and mortar. 5 g of
homogenate were mechanically stirred in 15 ml of a 4.5% (w/v)
solution of metaphosphoric acid for 15 min. The mixture was vacuum ltered and diluted with HPLC grade water; the total nal volume was measured and an aliquot was ltered through a 0.45 lm
lter prior to injection into the chromatographic column. The
instrumentation details were: HP Series 1100 (quaternary pump,
vacuum degasser, a Rheodyne 20 lL injection loop and a DiodeArray Detector, controlled by HPChemStation software); Hypersil
ODS column (250 * 4.6 mm) of particle size 5 lm; mobile phase:
HPLC grade water with metaphosphoric acid to pH 2.2; detection
at 245 nm; calibrated by external standard method.
2.3. Chlorophyll determination
Pigments were extracted from 100 mg of lyophilized spinach
with 50 mL of ice-cold aqueous acetone solution (80% v/v). Samples were homogenized with acetone for 2 min. All extraction procedures were performed in dim light and cold conditions. The
extracts were pooled and centrifuged at 9000g for 15 min at 4 C
(Olivera et al., 2008; Mazzeo et al., 2011). After the ltration of
extracts, a spectrometric system UVVis was used to determine
the concentration of the chlorophylls a and b in spinach. The measurement absorbance was conducted at 664 nm and 647 nm, at the
absorbance maxima for chlorophylls a and b in acetone (with 20%
water), respectively. These data were used to calculate total chlorophyll (Porra et al., 1989), chlorophyll a and chlorophyll b contents (Linchtenthaler and Buschmann, 2001).
2.4. Colour measurement
Colour of frozen spinach leaves was measured. Quantitation of
the colour change was based on measurement of CIELab values
(CIE, 1978) with a CR-200 Minolta Chromameter (Minolta Co.,
Chuo-Ku, Osaka, Japan). L indicates lightness, with values ranging
from 0 (completely opaque or black) to 100 (completely transparent or white); a positive a value indicates redness on the hue circle (a 1=4 greenness) and a positive b value indicates yellowness (b
1=
4 blueness). A standard white plate (Calibration plate CR-200) was
used to standardize the instrument under C illuminant condition
according to the CIE (Commission International de l Eclairage).
Eight to ten pieces of frozen leaves, representative of the products,
were measured at frequent time intervals during their storage at
isothermal and variable temperature conditions. Colour measurements were conducted for frozen spinach samples after cooking
(during the sensory evaluation, described below).
2.5. Texture measurement
The lm support rig HDP/FSR (containing perspex lm support
platform, aluminum circular top plate, thumb screws, 5 mm spherical probe) was used to measure the burst strength of spinach
leaves. Prior to performing the test, the sample was placed over a
hole in a raised Perspex platform. A top plate prevented the sample

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from slipping during testing. The test was then carried out as the
arm of the texture analyzer (MODEL TA-XT2i, Stable Micro Systems, Godalming, Surrey, U.K.), that brought a 5 mm stainless steel
ball probe down into the aperture. The maximum force to rupture
the product was recorded and was referred to as burst strength. For
each storage temperature, the burst strength was measured at two
different points of medium-sized non-injured spinach leaves (8
10) (at ambient temperature).

where CvitC and C0,vitC are the concentrations of L-ascorbic acid at

time t and zero respectively and kvitC is the apparent reaction of
vitamin C loss, estimated by the slope of the linearized plot of
ln(CvitC/C0,vitC) vs t. Arrhenius equation expressed adequately the
temperature dependence of reaction rate of vitamin C, kvitC (Eq. (2)):

kv itC

3
Ea
E
4
5 or ln kv itC ln kref ;v itC   a 
kref ;v itC exp  
1
1
1
R TT
R T  T1
ref

Sensory evaluation of the spinach samples was conducted by

10 panellists of the accredited (according to ISO 17025) sensory
laboratory of NTUA. The panellists were trained in the sensory
analysis (Lebesi and Tzia, 2011) and were experienced in assessing cooked vegetable products. Spinach samples (leaves) were
portioned (approximately 150 g), re-packed and tempered at
18 C for 24 h before their sensory assessment. The sensory
characteristics of frozen spinach were evaluated after cooking
(2 min in boiled water). Cooked spinach samples were provided
on white plastic plates coded with random 3-digit numbers, in
individual booths and evaluated twice. Every time, four samples
(1 control sample and 3 other samples) were evaluated, one
of which was the control. Overall visual acceptability, texture
(in mouth, expressed as juiceness), colour, aroma, avour/taste
and overall impression were rated based on the preference using
a 9-point hedonic scale (1: extremely dislike to 9: extremely like).
Freshness and off-odour development was rated. Rating was carried out using a nine point scale where 1 = absent or poor characteristic, 9 = too intense or excellent. Panellists were motivated to
express criticisms on the score sheets used for the sensory evaluation. The colour and textural attributes, also determined through
instrumental measurements, were included in the sensory analysis, so as to determine how these parameters affect the overall
acceptance of the product.
2.7. Statistical analysis
Duplicate measurements for each formulation were conducted.
The presented results are mean values of duplicate experiments
(n = 8 or 10 accordingly). The results were statistically analyzed
by analysis of variance (ANOVA) using Statistica 7.0 software
(StatSoft Inc., Tulsa, Okla., U.S.A.).
3. Results and discussion
3.1. Vitamin C loss in frozen spinach
The initial content of vitamin C in the frozen spinach leaves
studied was measured at 25.9 2.7 mg/100 g of frozen vegetable,
in agreement with the values reported for green fresh vegetables
(frozen broccoli 66.176.7 mg/100 g; frozen green beans 11.8
15.1 mg/100 g; frozen spinach 12.624.5 mg/100 g; frozen peas
19.621.8 mg/100 g) (Favell, 1998). The non-homogeneous nature
of raw material in spinach samples is reected in the signicant
variability of the initial nutritional value. This demonstrates the
need for caution in assuming a single xed value for the initial vitamin C content. The average retention of vitamin C is expressed relatively to an initial, average value of day 0 of the experiment,
where C represents the concentration of L-ascorbic acid in 100 g
of raw material. In all cases, vitamin C loss was found to be adequately described by an apparent rst order reaction (Eq. (1),
Table 1, Fig. 1):

C v itC C 0;v itC ekv itC t or ln

ref

2.6. Sensory evaluation

C v itC
kv itC t
C 0;v itC

where kref,vitC is the reaction rate of the vitamin C loss at a reference

temperature Tref (18 C), Ea is the activation energy of the chemical
reaction and R is the universal gas constant (Taoukis et al., 1997). By
linearly correlating ln kvitC vs (1/Tref  1/T) (Arrhenius plot), the Ea of
L-ascorbic oxidation was estimated from the slope of the tted line.
The estimated activation energies Ea, the 95% condence range as
well as the goodness of t (R2) for frozen spinach are shown in
Table 1. The activation energy Ea for vitamin C loss was calculated
as 132.0 5.8 kJ/mol. The high activation energy enables frozen
storage to kinetically constrain the L-ascorbic acid oxidation, which
makes frozen storage at low temperatures a good means of preserving vitamin C. However, this quality parameter is very sensitive to
abuse subzero storage temperatures and temperature uctuations
during the cold chain.
The differing behaviour regarding vitamin C oxidation of green
vegetables has been reported by Giannakourou and Taoukis (2003).
Vitamin C oxidation has been related to the different tissue structure, mechanical damage during harvesting, intrinsic enzyme
(ascorbate oxidase) and sulphydryl group content and the presence
of metal ions, such as Fe3+ and Cu2+ that act as catalysts. The loss of
ascorbic acid is probably enhanced by the activity of ascorbate oxidase, which is strongly dependent on the pH of the vegetable. The
spinach leaf is shown to be prone to lose rapidly its high initial
Vitamin C, due to the comparatively high value of surface area
and to an elevated amount of iron, that is known to play a decisive
role in the oxidative degradation of L-ascorbic acid to L-dehydroascorbic acid.
Vitamin C is widely used as an appropriate marker for monitoring quality changes during processing, storage and at the end point
of the frozen chain. This is a consequence of its concentration
change due to irreversible oxidation mechanisms that are
enhanced by temperature abuses (Serpen et al., 2007). In this
study, based on the results of preliminary experiments where Vitamin C loss and sensory testing were compared, 70% vitamin C loss
was used as the acceptability level for frozen spinach samples
(Table 6) in accordance with the level of sensory acceptability,
determined by the preliminary procedure. In published work,
lower values for vitamin C loss (e.g. 50%) had been used for the
shelf life calculation of vegetables (Giannakourou and Taoukis,
2003). Since the vitamin C loss follows 1st order kinetic, the shelf
life (SL) can be predicted according to Eq. (3). The shelf life is calculated as 23, 66, 100 and 401 days at storage temperatures 5,
8, 12 and 18 C, respectively (Table 6). Shelf life values from
earlier studies on the shelf life of frozen spinach can lead to different estimations, which may be attributed to different cultivars that
could affect the mode of deterioration, due to different enzyme
concentration, water activity and other factors.

ln
SL

C 0;v itC
C v itC

kv itC


3

The established kinetic model was validated at dynamic storage

conditions in programmable freezerincubator. To demonstrate
the integrated effect of the temperature variability on product
quality, the term of the effective temperature Teff is introduced
(Giannakourou and Taoukis, 2003). Teff, which is dened as the

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Table 1
Estimated kinetic parameters (the reaction rate of vitamin C loss, kvitC (1/d), the activation energy Ea (kJ/mol)) for vitamin C loss of frozen spinach leaves, at isothermal (5, 8,
12, 18 C) and non-isothermal timetemperature conditions (Teff = 6.9 C).
Kinetic parameters

Isothermal temperature conditions

Non-isothermal temperature conditions

5 C

8 C

12 C

18 C

6.9 C

kvitC (1/d)

0.0639 0.0061

0.0313 0.0027

0.0117 0.0009

0.0026 0.0002

kexp: 0.0351 0.0001

R2 for kexp: 0.999
keff,pre: 0.0344 0.0028

R2
Ea s.d. (kJ/mol)
kref (1/d) (Tref = 18 C)
R2

0.986
132.0 5.8
0.0029 0.0004
0.996

0.979

0.956

0.936

Timetemperature conditions of repeated cycles: 1: 12 h at 5 C; 2: 12 h at 8 C; 3: 12 h at 10 C, with calculated Teff value 6.9 C.
At T = 30 C, there was no difference observed in the vitamin C content.
s.d. standard deviation for two replicate determinations.
95% condence intervals based on the statistical variation of the kinetic parameters of the Arrhenius model.

where keff is the value of the rate of vitamin C loss at the effective
temperature. If the temperature prole is a step sequence, as in
our experiments, or is discretized to small time increments ti of constant temperature Ti, where Rti = ttot, then Eq. (4) can equivalently
be expressed as:

kv itC;ref



 
X
Ea 1
1
t i kv itC;eff ttot
exp 

R T i T ref
i

from which keff can be estimated. For k = keff, the value of the effective temperature Teff can be calculated from the Arrhenius equation
(Eq. (2)).
In Fig. 2, measurements of vitamin C loss and the corresponding
exponential t are shown and compared to predictions at the corresponding Teff, with the dotted lines representing the limits of 95%
condence range of the quality prediction for frozen spinach
leaves. Repeated temperature cycles included three step changes
as shown in Table 1 (1st step = 12 h at 5 C, 2nd step = 12 h at
8 C and 3rd step = 12 h at 10 C, Teff = 6.9 C). Predicted rate
of loss, keff,pre are in good agreement with the experimentally estimated ones, kexp, as it is demonstrated in Table 1. The two rates are
considered statistically equivalent in non-isothermal experiments.
Giannakourou and Taoukis (2003) reported shorter shelf-life for
frozen peas and spinach (25 and 5 days, respectively) exposed to
changeable temperatures (strongly depending on the vegetable

Fig. 1. (a) Results for vitamin C loss vs time at 4 storage temperatures.

Experimental points correspond to: 5 C (4), 8 C (h), 12 C (}), and 18 C
(s); lines represent the rst order t (Eq. (1)) (b) Arrhenius plot of the vitamin C
loss rate for frozen spinach (with Tref = 18 C) (Eq. (2)).

constant temperature that results in the same quality value as the

variable temperature distribution over the same time period, is
based on the Arrhenius model and integrates in a single value
the effect of the variable temperature prole. The nutritional
change under variable temperature conditions T(t) for time ttot
can be calculated by Eq. (4).


 Z ttot
C v itC;ttot

ln
kv itC Ttdt kv itC;eff ttot
C 0;v itC
0

Fig. 2. Comparison of experimental and predicted results of vitamin C loss of frozen

spinach leaves for exposure at the shown variable temperature prole. The solid
line represents the exponential t of the quality measurements and dotted lines
depict the upper and lower 95% condence range of quality predicted for Teff
(6.9 C).

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E. Dermesonluoglu et al. / Journal of Food Engineering 148 (2015) 1323

tissue). Frozen raspberry stored for 24 days under temperature

uctuations decrease c.a. 20% of its initial vitamin C content
(Sousa et al., 2005). However, Cruz et al. (2009) and Gormley
et al. (2002) did not notice variation in vitamin C content in watercress and strawberries respectively, submitted to time-varying
temperature conditions.

and experimental data. In all cases,

found to be adequately described by
(Fig. 3a) (Eq. (6)), as reported in the
1981; Schwartz and von Elbe, 1983;
and Tong, 1996; Koca et al., 2006):

C chl C 0;chl ekchl t or ln

3.2. Chlorophyll loss in frozen spinach
The colour of spinach is mainly related to chlorophyll content as
this compound is the main pigment of green vegetables and masks
the bright color of carotenoids. Chlorophylls a and b were detected
in frozen spinach samples with chlorophyll a being higher than
chlorophyll b exhibiting a chlorophyll a/chlorophyll b ratio 2.63
2.69. The main ratio of chlorophyll a to b was similar to the values
reported for other dark green leafy tissues. The initial chlorophyll
content of leafy spinach is measured as: chlorophyll a 37.4 3.4,
chlorophyll b 13.9 2.0 and total chlorophyll 50.8 4.7, mg/100 g
frozen vegetable (dry matter for spinach was calculated as
11.01 0.91 g/100 g.). Chlorophyll a and b degradation of spinach
leaves was observed during 250 days of frozen storage, respectively, for the 5 studied temperatures (isothermal and non-isothermal conditions). The loss of chlorophyll a and chlorophyll b with
storage time and temperature followed the same pattern. Total
chlorophyll content calculated was used as quality indicator,
exhibiting a good correlation coefcient between the tted model

total chlorophyll loss was

1st order reaction kinetics
literature (Schwartz et al.,
Canjura et al., 1991; Steet

C chl
kchl t
C 0;chl

where Cchl and C0,chl are the concentrations of total chlorophyll at

time t and zero respectively and kchl is the apparent reaction rate
of total chlorophyll loss, estimated by the slope of the linearized
plot of ln(Cchl/C0,chl) vs t (Table 2). The temperature dependence of
the deterioration rate k was then modeled by the Arrhenius equation (Fig. 3b) (Eq. (2)). The activation energy Ea for total chlorophyll
loss of frozen spinach was calculated as 70.3 11.3 kJ/mol. 90%
retention of chlorophyll has been reported as criteria for frozen
spinach shelf life determination in literature (Labuza, 1982; Reid
et al., 2003). More recently, 7080% chlorophyll retention has been
used for the quality estimation of frozen spinach and the investigation of the colour change (Philippon et al., 1986; Jaworska and
Kmiecik, 2000). In this study, the shelf life of frozen spinach was calculated based on 60% retention of total chlorophyll (Table 6). The
sensory limit of acceptance for frozen spinach samples were also
taken into account. The shelf life is calculated to be 27, 50, 119
and 464 days at storage temperatures 5, 8, 12 and 18 C,
respectively.
3.3. Colour deterioration in frozen spinach
Crucial to the attractive appearance of fresh and processed leaf
vegetables is the green colour, which depends on the amount of
chlorophylls and the ratio of chlorophyll a to chlorophyll b. Colour
loss during frozen storage is attributed to the fading of the vivid
green colour of chlorophyll to an olive brown, characteristic of pheophytin. Another common type of deterioration, is the removal of
the phytol chain, leading to the formation of chlorophyllide
(removal from chlorophyll) or pheophorbide (removal from pheophytin). Furthermore reactions related with the functional side
groups of chlorophyll from colourless end products also affect colour during frozen storage (e.g. the isocyclic ring may be oxidised to
form allomerised chlorophyll). Because pheophitisation reaction
rates are generally higher than other chlorophyll degradation pathways, it is considered the most important mechanism of chlorophyll destruction during food processing and at high storage
temperatures. However, the investigations by Jaworska and
Kmiecik (2000) on frozen spinach and New Zealand spinach and
by Philippon et al. (1986) on parsley shows a loss of chlorophylls
of 20% causes no, or only a slight change in the colour. Martins
and Silva (2002) also failed to nd any deterioration in colour in
frozen, unblanched French bean after 2 months frozen storage,
despite losses of chlorophyll of up to 89% (reported also by Reid
et al., 2003; Lisiewska et al., 2009). At lower storage temperatures,
colour is stabilised probably by the formation of metalchlorophyll
compounds. Therefore, although chlorophyll content and colour
deteriorate during frozen storage, and the chlorophyll content is
related to the initial vivid green colour, chlorophyll content does
not give a reliable prediction of colour retention.
Instrumentally measured spinach colour was expressed by the
value for total colour E according to Eq. (7).

E
Fig. 3. (a) Results for total chlorophyll content vs time at storage temperatures.
Experimental points correspond to: 5 C (4), 8 C (h), 12 C (}), 18 C (s)
and variable temperature conditions (O); lines represent the rst order t (Eq. (6))
(b) Arrhenius plot of the total chlorophyll loss rate for frozen spinach (with
Tref = 18 C) (Eq. (2)).

q
2
L2 a2 b

where L, a and b are colour parameters of the CIELab scale. The E

values of frozen spinach stored at isothermal temperature conditions 5, 8, 12 and 18 C and non-isothermal temperature conditions and times 0, 53, 102, 151, 200 and 250 days were presented

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E. Dermesonluoglu et al. / Journal of Food Engineering 148 (2015) 1323

Table 2
Estimated kinetic parameters (the reaction rate of total chlorophyll loss, kchl (1/d), the activation energy Ea (kJ/mol)) for total chlorophyll loss of frozen spinach leaves, at
isothermal (5, 8, 12, 18 C) and non-isothermal timetemperature conditions (Teff = 6.9 C).
Kinetic parameters

Isothermal temperature conditions

Non-isothermal temperature conditions

5 C

8 C

12 C

18 C

6.9 C

kchl (1/d)

0.0049 0.0003

0.0041 0.0002

0.0032 0.0002

0.0012 0.0002

kexp: 0.0040 R2 for kexp: 0.966

keff,pre: 0.0045

R2
Ea s.d. (kJ/mol)
kref (1/d) (Tref = 18 C)
R2

0.987
70.3 11.3
0.0011
0.944

0.991

0.984

0.803

Timetemperature conditions of repeated cycles: 1: 12 h at 5 C; 2: 12 h at 8 C; 3: 12 h at 10 C, with calculated Teff value 6.9 C.
At T = 30 C, there was no difference observed in the total chlorophyll.
s.d. standard deviation for two replicate determinations.
95% condence intervals based on the statistical variation of the kinetic parameters of the Arrhenius model.

in Fig. 4. Colour change was expressed as a decrease in the total

colour E. However, the obtained results for spinach colour change
during frozen storage could not be used for mathematical modelling
purposes. The non-homogeneous nature of raw material in spinach
samples was reected on the signicant variability of the initial
colour as well as the colour change during frozen storage.
3.4. Texture deterioration in frozen spinach
Burst strength values were plotted against storage time for each
storage temperature for the spinach leaves. The burst strength
(=maximum force to rupture) was found to increase when plotted
with time for each storage temperature studied. Lower storage
temperatures resulted in lower burst strength values when compared with corresponding values of spinach leaves stored at higher
temperatures. The increase of burst strength values F could be
described by a linear equation (Eq. (8), Fig. 5, Table 3).

F F o ktexture  t

where F is the burst strength value measured at each storage temperature at time t, F0 is the burst strength value for spinach leaves at
time t = 0 (F0 = 60) and ktexture is the increase burst strength value
constant rate for spinach leaves at each storage temperature. The

Fig. 5. Effect of storage time on the burst strength values (F) of frozen spinach
leaves, (a) experimental points correspond to: isothermal temperature conditions
5 C (4), 8 C (h), 12 C (}), 18 C (s) and variable temperature conditions
(O); lines represent the 0 order t and (b) Arrhenius plot of the burst strength
increase rate for frozen spinach (with Tref = 18 C).

Fig. 4. Calculated values for total colour (E) by instrumentally measured colour
parameters L, a, and b of frozen spinach leaves, at isothermal (5, 8, 12, 18 C)
and non-isothermal timetemperature conditions (Teff = 6.9 C).

results obtained are depicted in Fig. 5. In this study, the shelf life
of frozen spinach was calculated based on measured burst strength
value of 180 g as limit of acceptance for the texture parameter
(Table 6). The sensory limit of acceptance for frozen spinach samples obtained during sensory evaluation was used for setting the
burst strength value at the end of shelf life (limit of acceptance
for overall texture of frozen spinach samples).

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Table 3
Estimated kinetic parameters (the reaction rate of burst strength increase, ktexture (1/d), the activation energy Ea (kJ/mol)) for burst strength increase of frozen spinach leaves, at
isothermal (5, 8, 12, 18 C) and non-isothermal timetemperature conditions (Teff = 6.9 C).
Kinetic parameters

Isothermal temperature conditions

Non-isothermal temperature conditions

5 C

8 C

12 C

18 C

6.9 C

ktexture (1/d)

0.8232 0.1034

0.6046 0.1175

0.4235 0.0310

0.3017 0.0227

kexp: 0.6684 0.00710

R2 for kexp: 0.938
keff,pre: 0.6740

R2
Ea s.d. (kJ/mol)
kref (1/d) (Tref = 18 C)
R2

0.910
43.38 4.81
0.2872
0.976

0.794

0.968

0.954

Non-isothermal temperature conditions, timetemperature conditions of repeated cycles: 1: 12 h at 5 C; 2: 12 h at 8 C; 3: 12 h at 10 C, with calculated Teff value
6.9 C.
s.d. standard deviation for two replicate determinations.
95% condence intervals based on the statistical variation of the kinetic parameters of the Arrhenius model.

3.5. Sensory evaluation in frozen spinach

Scores for the sensory attributes evaluated were mathematically modelled by using apparent zero order kinetics for overall
visual acceptance, texture/juiceness, aroma, taste, freshness and
overall impression, and rst order kinetics for off-odour development (Eqs. (9) and (10). In Fig. 6, scores for the different sensory
attributes of cooked spinach leaves vs storage time at temperatures
studied were presented.

Ssensory

attribute

S0;sensory

attribute

 ksensory

attribute

Soff -odour S0;off -odour expkoff -odour  t

t

9
10

where S and So are the scores for the sensory attributes examined at
time t and zero respectively and ksensory attribute the apparent reaction
rate of the respective sensory attribute (Table 4). The temperature
dependence of the deterioration rate ksensory attribute was then modeled by the Arrhenius equation (Fig. 7, Table 5). The activation
energy Ea for the different sensory attributes of frozen spinach
was calculated from 29.21 (aroma) to 65.65 (freshness) kJ/mol.
The respective value for the effect of the storage temperature on
the overall impression of frozen spinach was 61.26 3.36 kJ/mol.
The score 5 (Sf), for overall impression was used as the acceptability level for frozen leafy spinach samples based on the rejection
response of panellists that rated on the hedonic scale. Considering
the apparent zero order kinetic, the shelf life (SL) can be thus calculated by (Eq. (11)):

SL

S0;ov erall

impression

kov erall

 Sf ;ov erall

impression

11

impression

In Table 6, calculated shelf-life values for frozen leafy spinach

samples stored at different temperatures are presented. Eq. (11)
is a simple mathematical tool allowing calculation of the shelf life
of frozen spinach accounting for the overall sensory acceptability.
The shelf life is calculated as 148, 195, 283 and 506 days at storage
temperatures 5, 8, 12 and 18 C, respectively. Reid et al.
(2003) also reported that the practical storage life for frozen spinach was 110 and 210 days at temperatures 6.7 and 12.3 C,
respectively.
In Table 6 calculated values for shelf life of frozen leafy spinach
samples, based on different quality indicators (70% vitamin C loss,
60% total chlorophyll retention, score for acceptability 5/9 during
sensory evaluation), are presented. Nutritional (Vitamin C) degradation is an important indirect quality index, but the fact that commercial and consumer acceptability are mainly based on sensory
quality criteria, such as colour, aroma, avour, texture and juiceness should not be overlooked. In case of nutritional labeling,

however, shelf life dating is legally limited by nutrient declaration and thus, can become the shelf life determining criterion. In
all cases, vitamin content can be used as a reliable index that
reects the whole temperature history of the product. Validated
kinetic models of vitamin C can be used for evaluation, control
and proper management of the frozen chain. Instrumentally measured texture was modelled and calculated shelf life values based
on texture where in good agreement with ones calculated from
sensory acceptability. Instrumentally measured colour could not
be mathematically modeled.
In order to demonstrate the usefulness of kinetic models, realistic timetemperature scenarios are considered, based on recent
surveys of the actual frozen distribution chain. The importance of
this validation is sustained by the diversity of timetemperature
handling of a product during its actual distribution in the post-processing chain, to the consumer end (Giannakourou and Taoukis,
2007) as conrmed by recent data, retrieved by the user-friendly
database of the FRISBEE project (Food Refrigeration Innovations
for Safety, Consumers Benet, Environmental Impact and Energy
Optimisation Along the Cold Chain in Europe, FP7, www.frisbeeproject.eu/Data available at: www.frisbee-project.eu/coldchaindb.html). Based on this set of data, representing more than 200
consumer freezers, in different countries it was shown that in
almost 30% of consumer storage the temperature was higher than
14 C, with 10% exceeding the 10 C. On the other hand, a significant percentage of about 9.5% was measured stored at very low
temperatures (below 22 C) (Fig. 8a). As far as retail display storage is concerned, from data from the same database (including
almost 320 cases), more than 60% was found to operate at temperatures at or below 18 C to preserve frozen foods, and only 1.5
was measured to have average temperatures higher than 14 C
(Fig. 8b).
To fully understand the need to take into account the real time
temperature history of the product, and not assume a common scenario, two alternative scenarios were applied in the case of frozen
spinach. Therefore, using Eqs. (4) and (5), and the kinetic results of
Tables 1 and 4 (concerning vitamin C and overall sensory impression respectively), the shelf life consumed at the end of each stage
was estimated. To illustrate the importance of assessing the effect
of the real temperature conditions during transport and storage on
food quality, a realistic scenario of 240 days distribution was studied, based on FRISBEE database. It included an initial stage of
40 days storage in the packing plant, followed by transportation
and storage in a distribution center for 40 days. Subsequently,
packages of frozen spinach were kept at the retail display for
60 days, before being purchased by the nal consumers that store
them in their domestic freezer for 100 days before consumption. At
the end of the assumed cycle, i.e. the time of consumption, the

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E. Dermesonluoglu et al. / Journal of Food Engineering 148 (2015) 1323

Fig. 6. Scores for different sensory attributes of cooked spinach leaves vs time at storage temperatures studied. Experimental points correspond to: isothermal temperature
conditions 5 C (4), 8 C (h), 12 C (}), 18 C (s) and variable temperature conditions (O); lines represent the 0 (for all sensory attributes) or 1st order t (for offodour): (a) overall visual acceptability, (b) colour, (c) off-odour, (d) texturejuiceness, (e) aroma, (f) taste, (g) overall acceptability and (h) freshness.

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E. Dermesonluoglu et al. / Journal of Food Engineering 148 (2015) 1323

Table 4
Estimated kinetic parameters used for quality deterioration of frozen spinach leaves stored at isothermal (5, 8, 12 and 18 C) and non-isothermal timetemperature
conditions (Teff = 6.9 C) based on scoring (on 19 scale) of different sensory attributes (off-odour, overall visual acceptability, texturejuiceness, aroma, taste, freshness and
overall acceptability).
Sensory attribute

Kinetic parameters

5 C

8 C

12 C

18 C

Non-isothermal temperature
conditions
6.9 C

Off-odour

S0
k (1/d)
R2

1.33 0.190
0.0107 0.0002
0.974

0.0066 0.0002
0.956

0.0041 0.0001
0.952

0.0010 0.0003

0.0070 0.000
0.988

S0
k (1/d)
R2

8.71 0.08
0.0239 0.0022
0.942

0.0129 0.0005
0.988

0.0115 0.0009
0.951

0.0068 0.0010
0.873

0.0172 0.0008
0.984

S0
k (1/d)
R2

8.58 0.08
0.0235 0.0018
0.949

0.0140 0.001
0.943

0.0105 0.0009
0.950

0.0062 0.0010
0.868

0.0167 0.0020
0.927

S0
k (1/d)
R2

8.75 0.08
0.0269 0.0010
0.964

0.0168 0.0004
0.986

0.0132 0.0003
0.986

0.0091 0.0008
0.888

0.018 0.0005
0.984

S0
k (1/d)
R2

8.88 0.07
0.0286 0.0012
0.965

0.0178 0.0003
0.993

0.0141 0.0003
0.988

0.0050 0.0003
0.934

0.0194 0.0004
0.990

S0
k (1/d)
R2

8.88 0.07
0.0282 0.0012
0.958

0.0173 0.0006
0.969

0.0136 0.0003
0.988

0.0051 0.0004
0.878

0.0183 0.0005
0.983

S0
k (1/d)
R2

8.84.0.05
0.0287 0.008
0.980

0.0189 0.0004
0.995

0.0133 0.0003
0.984

0.0071 0.0003
0.975

0.0200 0.0003
0.990

Overall visual acceptability

Texturejuiceness

Aroma

Taste

Freshness

Overall impression

Isothermal temperature conditions

Timetemperature conditions of repeated cycles: 1: 12 h at 5 C; 2: 12 h at 8 C; 3: 12 h at 10 C, with calculated Teff value 6.9 C. Results were referred to experimental
values (kexp, R2 for kexp).
s.d. standard deviation for two replicate determinations.
95% condence intervals based on the statistical variation of the kinetic parameters of the Arrhenius model.

Table 5
Arrhenius equation parameters (Ea and kref,sensory attribute) and statistics (R2) for quality
loss of frozen leafy spinach samples (Temperature range = 18 to 5 C, Tref = 18 C)
based on scoring (on 19 scale) of different sensory attributes (off-odour, overall
visual acceptability, texturejuiceness, aroma, taste, freshness and overall
acceptability).

Fig. 7. Arrhenius plot of the quality loss rate for frozen spinach based on different
sensory attributes evaluated, overall visual acceptability (), off-odour (d), texture
juiceness (s), aroma (), taste (N), freshness (.) and overall impression (h)
(Tref = 18 C).

remaining shelf life of frozen spinach (at 18 C) according to vitamin C and overall impression values was estimated.
The rst scenario of 240 days distribution cycle, scenario (a)
(Fig. 9) was one, with very low temperatures at the rst two stages
of the chain (warehouse and distribution center) and appropriate
storage conditions during retail display and consumer storage (corresponding to almost 40% of Fig. 8a). The second scenario (b)
(Fig. 9) was the same one, regarding the rst three stages, but took
into account the possibility of abusive storage at the nal crucial

Sensory attribute

kref,sensory attribute
(1/d) s.d.

Ea (kJ/mol)

R2

Off-odour
Colour
Overall visual acceptability
Texturejuiceness
Aroma
Taste
Freshness
Overall impression

0.0026 0.0008

0.0060 0.0010
0.0050 0.0010
0.0080 0.0010
0.0063 0.0016
0.0058 0.0016
0.0077 0.0007

43.49 10.6

53.42 5.76
62.52 5.70
47.71 4.18
63.66 3.75
64.98 4.12
61.26 3.36

0.881

0.998
0.998
0.999
0.999
0.951
0.988

stage of domestic storage (corresponding to almost 10% of

Fig. 8a). In Fig. 10, remaining shelf life (estimated assuming an isothermal storage at 18 C) was plotted at the end of each stage for
both scenarios. It was observed that, for scenario (a), at the end of
the cycle, the frozen product was still of more than acceptable
quality, based on both criteria (30 days and 200 days for vitamin
C and sensory scoring respectively of remaining shelf life if stored
at 18 C for the remaining time). On the other hand, for the abusive scenario (b), the product reached the threshold of vitamin C
loss a month after its storage in the domestic freezer but was still
sensorially of good quality (60 days of remaining shelf life)
(Fig. 10).
Based on the aforementioned observations, the signicance of
monitoring the real timetemperature history of the product
throughout its distribution was illustrated, in order to reliably
estimate its quality status, at any point of its lifecycle. If the

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E. Dermesonluoglu et al. / Journal of Food Engineering 148 (2015) 1323

Table 6
Shelf life (SL, days) calculation for frozen leafy spinach samples based on different quality indicators (70% vitamin C loss, 60% total chlorophyll retention, score for acceptability 5/9
during sensory evaluation, nal value for burst strength 180) (temperature range = 20 to 5 C).
Criterion

70% vitamin C loss

60% total chlorophyll content retention
Texture (Final value for burst strength = 180)
Sensory evaluation (Final score for overall impression = 5/9)

Shelf life (SL, days) calculation for leafy frozen spinach

5 C

8 C

12 C

18 C

20 C

23
27
155
148

66
50
193
195

100
119
261
283

401
464
418
506

641
740
491
618

Fig. 10. Remaining shelf life of frozen spinach at the end of each stage of the cold
chain, for scenario a and b.

4. Conclusions

Fig. 8. Temperature distribution in: (a) domestic freezers and (b) retail display.
Source: database of FRISBEE project (2013).

Based on the obtained results, chlorophyll degradation, texture

measured instrumentally as rupture resistance and overall sensory
impression of frozen spinach leaves can be used as reliable quality
indices correlating to the products end of shelf life. Validated
kinetic models of these quality attributes can be used for evaluation, control and proper management of the frozen chain, as well
as for predicting product quality status at any stage of its distribution. If, however, nutritional labelling is applied, one must reconsider the application of nutritional criteria, such as vitamin C loss
for the estimation of their shelf life. According to results the shelf
life values calculated based on sensory evaluation were longer than
the respective values estimated by the kinetic modelling of the
vitamin C loss at temperatures higher than 18 C (at T = 18 C
based on sensory evaluation and vitamin C loss, 506 and 401 days,
respectively). Therefore, it becomes apparent that it is important to
identify the shelf life determining factors for specic frozen vegetables, since this decision leads to signicantly different
estimations.
Acknowledgements

Fig. 9. Cold chain conditions for frozen spinach. Source: FRISBEE project database
(2013).

temperature conditions of the products could be continuously

monitored, e.g. by inexpensive Time Temperature Integrators
(TTI), reliable estimation of the quality status and the remaining
shelf life of the products could be performed based on the presented modelling of the quality indices. This could allow better
management and optimization of the cold chain from manufacture
to consumption.

The research leading to these results has received funding from

the European Communitys Seventh Framework Programme (FP7/
2007-2013), FRISBEE (Food Refrigeration Innovations for Safety,
consumers Benet, Environmental impact and Energy optimisation along the cold chain in Europe) under Grant Agreement No.
245288 (http://www.frisbee-project.eu/).
References
Bunea, A., Andjelkovic, M., Socaciu, C., Bobis, O., Neascu, M., Verhe, R., Van Camp, J.,
2008. Total and individual carotenoids and phenolic acids content in fresh,
refrigerated and processed spinach (Spinaciaoleracea L.). Food Chem. 108, 649
656.

E. Dermesonluoglu et al. / Journal of Food Engineering 148 (2015) 1323

Canjura, F.L., Schwartz, S.J., Nunes, R.V., 1991. Degradation kinetics of chlorophylls
and chlorophyllides. J. Food Sci. 56, 16391643.
CIE, 1978. Recommendations on uniform color spaces color difference equations,
psychometric color terms. Supplement No 2 to CIE Publication No 15 (E-1.3.1)
1971/(TC-1-3), Commission Internationale de l Eclairage. Paris.
Cruz, R.M.S., Vieira, M.C., Silva, C.L.M., 2009. Effect of cold chain temperature abuses
on the quality of frozen watercress (Nasturtium ofcinale R. Br.). J. Food Eng. 94,
9097.
EE 1080/94/000069, 1995. Report from the research on the temperatures of frozen
products.
Favell, D.J., 1998. A comparison of the vitamin C content of fresh and vegetables.
Food Chem. 62 (1), 5964.
Giannakourou, M.C., Taoukis, P.S., 2002. Systematic application of time temperature
integrators as tools for control of frozen vegetable quality. J. Food Sci. 67 (6),
22212228.
Giannakourou, M.C., Taoukis, P.S., 2003. Kinetic modelling of vitamin C loss in
frozen green vegetables under variable storage conditions. Food Chem. 83, 33
41.
Giannakourou, M.C., Taoukis, P.S., 2007. Reaction kinetics. In: Sablani, S., Rahman,
M.S., Datta, A., Mujumdar, A.S. (Eds.), Handbook of Food and Bioprocess
Modeling. Marcel Dekker Inc., New York.
Gormley, R., Walshe, T., Hussey, K., Butler, F., 2002. The effect of uctuating vs.
constant frozen storage temperature regimes on some quality parameters of
selected food products. Lebensmittel.-Wiss. Technol. 35, 190200.
International Institute of Refrigeration, IIR, 1986. IIR. In: Recommendations for the
Processing and Handling of Frozen Foods Paris.
Jaworska, G., 2005. Nitrates, nitrites, and oxalates in products of spinach and New
Zealand spinach. Effect of technological measures and storage time on the level
of nitrates, nitrites, and oxalates in frozen and canned products of spinach and
New Zealand spinach. Food Chem. 93, 395401.
Jaworska, G., Kmiecik, W., 2000. Comparison of the nutritive value of frozen spinach
and New Zealand spinach. Pol. J. Food Nutr. Sci. 9/50 (4), 7984.
Kmiecik, W., Lisiewska, Z., 1999. Effect of pretreatment and conditions and period of
storage on some quality indices of frozen chive (Allium schoenoprasum L.). Food
Chem. 67, 6166.
Koca, N., Karadeniz, F., Burdurlu, H.S., 2006. Effect of pH on chlorophyll degradation
and colour loss in blanched green peas. Food Chem. 100, 609615.
Labib, A.S.S., Abd El-Latife, S.A., Omran, H., 1997. Quality indices of Jews mallow
and spinach during frozen storage. Plant Foods Hum. Nutr. 50, 333347.
Labuza, T.P., 1982. Shelf-life Dating of Foods. Food & Nutrition Press, Inc., Westport,
Connecticut 06880 USA.
Lebesi, D.M., Tzia, C., 2011. Staling of cereal bran enriched cakes and the effect of an
ndoxylanase enzyme on the physicochemical and sensorial characteristics. J.
Food Sci. 76 (6), S380-7.
Linchtenthaler, H.K., Buschmann, C., 2001. Chlorophylls and carotenoids Measurement and characterisation by UVVIS. In: Current Protocols in Food
Analytical Chemistry, John Wiley&Sons, Madison, pp. F4.3.1-F4.3.8.
Lisiewska, Z., Gebczynski, P., Bernas, E., Kmiecik, W., 2009. Retention of mineral
constituents in frozen leafy vegetables prepared for consumption. J. Food
Compos. Anal. 22, 218223.

23

Martins, R.C., Silva, C.L.M., 1998. Colour and chlorophylls degradation kinetics of
frozen green beans (Phaseolus vulgaris L.). In: Proc. of the 3rd International
Conference on Predictive Modelling in Foods, Wageningen, The Netherlands, pp.
185187.
Martins, R.C., Silva, C.L.M., 2002. Modelling colour and chlorophyll losses of frozen
green beans (Phaseolus vulgaris L.). Int. J. Refrig 25 (7), 966974.
Mazzeo, T., NDri, D., Chiavaro, E., Visconti, A., Fogliano, V., Pellegrini, N., 2011. Effect
of two cooking procedures on phytochemical compounds, total antioxidant
capacity and colour of selected frozen vegetables. Food Chem. 128, 627633.
Olivera, D.F., Via, S.Z., Marani, M., Ferreyra, R.M., Mugridge, A., Chaves, A.R.,
Mascheroni, R.H., 2008. Effect of blanching on the quality of Brussels sprouts
(Brassica oleracea L. gemmifera DC) after frozen storage. J. Food Eng. 84, 148
155.
Philippon, J., Rouet-Mayer, M.A., Fontenay, P., Duminil, J.M., 1986. Storage time and
temperature in relation to stability of chlorophylls, colour, and total release of
volatiles in frozen parsley. Sci. Aliment. 3, 433446.
Porra, R.J., Thompson, W.A., Kriedemann, P.E., 1989. Determination of accurate
extinction coefcients and simultaneous equations for assaying chlorophylls a
and b extracted with four different solvents: verication of the concentration of
chlorophyll standards by atomic absorption spectroscopy. Biochim. Biophys.
975, 384394.
Pukszta, T., Palich, P., 2007. The effect of freezing conditions of strawberry storage
on the level of thawing drip loss. Acta Agrophys. 9, 203208.
Reid, D.S., Kotte, K., Kilmartin, P., Young, M., 2003. A new method for accelerated
shelf-life prediction for frozen foods. J. Sci. Food Agric. 83, 10181020.
Schwartz, S.J., von Elbe, J.H., 1983. Kinetics of chlorophyll degradation to
pyropheophytin in vegetables. J. Food Sci. 48, 13031306.
Schwartz, S.J., Woo, S.L., von Elbe, J.H., 1981. High performance liquid
chromatography of chlorophylls and their derivatives in fresh and processed
spinach. J. Agric. Food Chem. 29, 533535.
Serpen, A., Gokmen, V., Bahceci, K.S., Acar, J., 2007. Reversible degradation kinetics
of vitamin C in peas during frozen storage. Eur. Food Res. Technol. 224, 749
753.
Sousa, M.B., Canet, W., Alvarez, M.D., Tortosa, M.E., 2005. The effect of the pretreatments and the long and short-term frozen storage on the quality of
raspberry (cv. Heritage). Eur. Food Res. Technol. 221, 132144.
Steet, J.A., Tong, C.H., 1996. Degradation kinetics of green colour and chlorophylls in
peas by colorimetry and HPLC. J. Food Sci. 61, 924927.
Taoukis, P.S., Fu, B., Labuza, T.P., 1991. Timetemperature indicators. Food Technol.
45, 7082.
Taoukis, P., Labuza, T.P., Sagy, S., 1997. Kinetics of food deterioration and shelf life
prediction. In: Sagy, S., Rotstein, K. (Eds.), Food Engineering Handbook. CRC
Press, Denver, CO, pp. 363405, Chapter 9.
Van Buggenhout, S., Lille, M., Messagie, I., Van Loey, A., Autio, K., Hendrickx, M.,
2006. Impact of pretreatment and freezing conditions on the microstructure of
frozen carrots: qualication and relation to texture loss. Eur. Food Res. Technol.
222, 543553.