CBC: Combined blast and contact cooling, RTS: Real temperature simulation LC: Liquid cooling, NC: No cooling, GP: Gel pack, S: Storage at -1 °C .... Average Torry freshness scores CBC: C
Experimental design
Cod used in the experiments (I and II) was trawler caught east of Iceland on February
On 22nd, 2009 (average ambient air temperature 5 °C), cod were bled, gutted and washed, then stored in crushed plate ice (fish-to-ice ratio approx 3:1) in 460 L tubs on board vessels for transport to a main processing facility in northern Iceland At the processing plant, the cod were processed in different ways on February 24 (11:00–14:00), with the average temperature of the fish during processing at 1 °C The cod loins were packed in 5 kg expanded polystyrene (EPS) boxes, which were palletized and containerized The procedures for experiments I and II followed.
Experiment I involved transporting pallets to Reykjavik in a refrigerated container from 5:00 pm to midnight, arriving at Matís, Reykjavík, at approximately 9:30 am the following day At Matís, the loins were stored for up to 13 days from processing, either at -1 °C or under real air freight temperature simulation (RTS).
The experimental groups were as follows:
A Liquid cooling (LC) and CBC cooling, with a cooling gel pack (GP) and stored under RTS conditions at Matís
B Liquid cooling and CBC cooling, stored under RTS conditions at Matís
C Liquid cooling, with a cooling gel pack and stored under RTS conditions at Matís
D No cooling (NC) during processing, with a cooling gel pack and stored under RTS conditions at Matís
E Liquid cooling and CBC cooling, with a cooling gel pack and stored at -1 °C (S) at Matís
F Liquid cooling and CBC cooling, stored at -1 °C at Matís
Following abbreviations of experimental groups will be used hereafter:
Sampling was done regularly from processing up to 13 days as shown in Table 1
Table 1 documents the sampling days used in Experiment I and lists the cooling conditions tested: CBC (combined blast and contact cooling), RTS (real temperature simulation during storage), GP (gel pack in boxes), LC (liquid cooling only), NC (no cooling during processing), and S (superchilled during storage).
Group Treatment Storage Sampling days*
CBC-RTS-GP CBC cooling RTS, gel pack 0, 1, 3, 6, 8, 10
CBC-RTS CBC cooling RTS, no gel pack 0, 1, 3, 6, 8, 10
LC-RTS-GP Liquid cooling RTS, gel pack 0, 1, 3, 6, 8, 10
NC-RTS-GP No cooling RTS, gel pack 0, 1, 3, 6, 8, 10
CBC-S-GP CBC cooling -1 °C, gel pack 0, 2, 7, 9, 13
CBC-S CBC cooling -1 °C, no gel pack 0, 2, 7, 9, 13
*Sampling on day 0 was done for microbial and chemical analysis only
Experiment II examined a subset of the cod loins lot described above, including CBC-cooled loins with and without gel packs After packaging in EPS boxes, these samples were transported to Bremerhaven, Germany by both air and sea freight to evaluate packaging and transport performance.
The cod loins were transported from the pick-up location in Bremerhaven/Cuxhaven to TTZ for temperature check, collection of data loggers and repackaging as retail portions
Two loins per package, with an average loin weight of 312 g, were stored under a modified atmosphere (MA) at 1 °C for up to 21 days from processing in Iceland, and during storage, samples were analysed by microbial and chemical methods at TTZ in Bremerhaven, Germany.
Fish loins were divided into eight experimental groups to assess the effects of transport mode (air or sea), packaging conditions (with or without a gel pack), and packaging method (MAP or air) Sampling was performed at arrival—two days for air freight and six days for sea freight from processing—and subsequent sampling after repackaging occurred at 3, 6, 11, 14 and 18 days for air-freight fish, with the corresponding sea-freight time points partially provided in the excerpt.
Sea freight fish were sampled at 10 and 15 days from repackaging For air freight fish, the last sampling day involved examining only MAP samples The last sampling day was, in fact, 20.
21 days from processing for air and sea freight fish, respectively
Table 2 Sampling groups in Experiment II
Air freight With gel pack FG MAP FG AIR
Without gel pack GG MAP GG AIR
Sea freight With gel pack FS MAP FS AIR
Without gel pack GS MAP GS AIR
Figure 1a Packaging of fish loins Figure 1b Gas composition of modified atmosphere packages
Upon arrival, the fish were repackaged in two formats: modified atmosphere packaging (MAP) with a gas composition of 45% N2, 5% O2, and 50% CO2, and conventional air packaging Each bag contained two cod loins stored at 1 °C in a cold chamber A data logger placed with the fish recorded an average temperature of 1.1 ± 0.6 °C, with the high standard deviation attributed to the cold chamber’s large doors being opened during sampling.
Temperature and humidity measurements
Experiments I and II used two types of loggers along with a relative humidity–temperature logger to record temperature and humidity The iButton temperature loggers (Figure 2), specifically the DS1922L model from Maxim Integrated, were employed for temperature measurements This device provides an accuracy of ±0.5 °C, a resolution of 0.0625 °C, and an operating range from -40 °C to 85 °C.
The experiments used iButton temperature loggers with dimensions of 17 mm in length and 5 mm in thickness to monitor product temperature To prevent microbial contamination, the loggers were placed inside plastic bags These measurements were carried out with Onset temperature loggers, type UTBI-001.
UTBI-001 data logger delivers accurate ambient temperature measurements with ±0.2°C accuracy, 0.02°C resolution, and an operating range of -20°C to 70°C Its compact dimensions—30 mm in diameter and 17 mm thick—make it suitable for use in climate chambers, where reliable ambient temperature data is essential for performance validation and process monitoring.
Figure 2.a iButton temperature logger Figure 2.b Onset Tidbit temperature logger c UB12 HOBO® Relative humidity and temperature data logger
Time accuracy: ± 1 minute per week
Four iButton temperature loggers were placed inside each EPS box and two on the outside of each box to monitor internal and external temperatures In Experiment 1, five Onset temperature loggers were used in each climate chamber to measure ambient temperature The configuration of temperature loggers for Experiment II is described in Section 4.1.
Figure 3 illustrates the locations of the temperature loggers inside the boxes, using a numbering scheme that is consistent with Table 3, and the logger names described hereafter correspond to those displayed in Table 3.
Figure 3 Location of temperature loggers inside each box The fish pile thickness in the box is represented by h and the vertical position of the loggers by z
Table 3 Location and numbering scheme of temperature loggers
Sensory evaluation
Experiment I Quantitative Descriptive Analysis (QDA), introduced by Stone and Sidel
Cooked cod samples (MA09sky027-040) were assessed using the Torry freshness score sheet (Shewan et al., 1953) and a 2004 evaluation protocol (Table 1) Eleven panellists trained according to ISO 8586 (1993) participated, with prior experience in cod sensory analysis and familiarity with QDA; they completed one training session before testing and used an unstructured 0–100 scale to rate the intensity of 30 sensory attributes described in Table 4 (attributes partly defined in Sveinsdottir et al., 2009; Magnússon et al., 2006) From the loins, samples about 40 g were placed in aluminum boxes coded with three-digit random numbers They were cooked for 6 minutes in a pre-warmed Convotherm oven at 95–100°C with air circulation and steam, then served to the panel Each panellist evaluated duplicates of each sample in random order across 14 sessions (no more than four samples per session), and data were recorded with a computerized system (FIZZ, Version 2.0, Biosystèmes, 1994–2000).
Data analysis included Principal Component Analysis (PCA) on the significant mean values of QDA sensory attributes with full cross-validation Analysis of Variance (ANOVA) was performed on the QDA data using the NCSS 2000 statistical program (NCSS, Utah, USA) Post hoc comparisons were conducted with Duncan’s multiple range test in NCSS, and the significance level was set at 5% unless stated otherwise.
Table 4 Sensory vocabulary for cooked samples of cod (Gadus morhua)
Key sensory attributes and their odors are as follows: o-sweet (a sweet odor); o-shellfish (shellfish and algae aromas characteristic of fresh seafood); o-meat (a meaty aroma reminiscent of boiled meat or halibut); o-vanilla (vanilla and warm milk notes); o-potatoes (an odor reminiscent of boiled potatoes); o-frozen (odors associated with refrigerator or freezer storage); and o-cloth (a damp cloth smell akin to a table cloth that has been left for about 36 hours).
Cod quality is assessed by sensory cues including TMA odour, which resembles dried salted fish and amine sour notes, and sour odours similar to spoiled milk or acetic acid with possible sulphur hints Visual cues show a contrast between ends: the left end should be light, white, and homogeneous, while the right end may be dark, yellowish, brownish, or grey and discoloured with stains; white precipitation on the surface may also appear In taste, a salty flavour and a metallic tang are common, with a characteristic metallic flavour often noted in fresh cod; a sweet flavour typical of fresh boiled cod, a meaty flavour reminiscent of boiled meat, and flavours linked to frozen storage, such as a wood-like note from refrigerator/freezer exposure, may be detected Pungent notes and sour flavours signal spoilage.
Evaluation centers on TMA (trimethylamine) and its f‑TMA amine off-flavour, which evokes a dried salted fish note and signals spoilage flavour intensity Off-flavour strength is measured alongside textural cues; the fish portion shows t-flakes as it flakes apart when pressed with a fork Texture varies along the length: the left end is firm, the right end soft, and overall firmness is assessed at the first bite Juiciness (t-juicy) is observed, with the left end tending to be dry This sensory profile—TMA flavour, off-flavour intensity, flakiness, asymmetric firmness, and juiciness—provides a clear, SEO-friendly description for quality assessment and product optimization.
Texture evaluation shows the right end is juicy, but after chewing several times it reads dry as it draws juice from the mouth, yet remains tender (t-tender) The left end comes off as tough, while the right end stays tender after repeated chewing With further chewing, mushiness emerges (mushy t-mushy), giving a mushy texture and meaty mouthfeel (t-meaty) that reflects a true meaty texture with crude muscle fibers A clammy sensation (clammy t-clammy) adds a clammy texture, and when tannins from a dry red wine are present, the bite becomes rubbery (t-rubbery) and springy.
Microbial measurements
Experiment I quantified total viable psychrotrophic counts (TVC) and counts of H2S-producing bacteria on iron agar (IA) following Gram et al (1987), but with 1% NaCl instead of 0.5% and without an overlay; plates were incubated at 17 °C for 5 days, and black colonies on IA indicate H2S production from sodium thiosulfate and/or cysteine For presumptive pseudomonads, Cephaloridine Fucidin Cetrimide (CFC) agar was modified according to Stanbridge and Board (1994) and used, along with Pseudomonas Agar Base (Oxoid) with CFC Selective Agar Supplement, with plates incubated at 22 °C for 3 days, and Pseudomonas spp form pink colonies on this medium Counts of Photobacterium phosphoreum were estimated by the method described.
13 using the PPDM-Malthus conductance method (Dalgaard and others 1996), as described by Lauzon (2003)
In all experiments, cooled Maximum Recovery Diluent (MRD, Oxoid) was used for dilutions, and agar media were surface-plated All samples were analyzed in triplicate, and results were presented as averages.
Experiment II, conducted at TTZ, Bremerhaven, Germany, quantified microbial populations by counting total viable counts (TVC) and H2S‑producing bacteria on Triple Sugar Iron Agar (Merck) incubated at 15 °C for 5 days, and by counting pseudomonads on CFC medium (Oxoid) incubated at 25 °C for 3 days In all experiments, cooled MRD was used for dilutions and agar media were surface‑plated All samples were analysed in duplicate and results presented as averages Counts of TVC, H2S‑producing bacteria and pseudomonads were reported as mean values.
Photobacterium phosphoreum were quantified by a quantitative PCR approach One milliliter of tenfold-diluted fish samples in MRD buffer was stored at -20 °C for subsequent DNA extraction For DNA isolation, the diluted samples were centrifuged at 11,000 × g for 7 minutes to form a pellet, the supernatant discarded, and DNA recovered from the pellet using the Promega Magnesil KF Genomic DNA Isolation Kit (MD1460) in conjunction with the KingFisher magnetic-bead automated DNA isolation instrument (Thermo Labsystems) following the manufacturers’ recommendations.
PCR amplification was performed on the Mx3005p instrument using Brilliant QPCR Mastermix (Stratagene, La Jolla, CA, USA) Primers were synthesized and purified by HPLC (MWG, Ebersberg, Germany) The DNA standard used for quantification was calibrated against the PPDM-Malthus conductance method.
Chemical analysis
Total Volatile Base Nitrogen (TVB-N) and Trimethylamine (TMA)
Experiment I used the Malle and Tao (1987) protocol to measure total volatile bases (TVB-N) and trimethylamine (TMA) TVB-N was quantified by steam distillation using a Struer TVN distillatory (STRUERS, Copenhagen) followed by titration, after extracting the fish muscle with 7.5% aqueous trichloroacetic acid (TCA).
Total volatile base nitrogen (TVB-N) was distilled and collected in a boric acid solution, then titrated with sulfuric acid Trimethylamine (TMA) was measured in a trichloroacetic acid (TCA) extract by adding 20 mL of 35% formaldehyde, which binds mono- and diamines under alkaline conditions, with TMA identified as the only volatile and measurable amine All chemical analyses were performed in triplicate.
Experiment II involved measuring Total Volatile Base Nitrogen (TVB-N) at two to three sampling points, in duplicate, by direct steam distillation into boric acid using a Kjeldahl-type distillatory, in accordance with German Food Law (LMBG) pH measurements were also taken.
In Experiment I, pH was measured in a mixture of 5 g minced loins and 5 mL deionised water using a Radiometer PHM 80 pH meter, calibrated at 25 °C with buffer solutions of pH 7.00 ± 0.01 and 4.01 ± 0.01 (Radiometer Analytical A/S, Bagsvaerd, Denmark).
Experiment II pH measured accordingly as above
Experiment I The water content of each loin was measured by accurately weighing out 5 grams of the minced sample in a ceramic bowl with sand The sample was then mixed to the sand and dried in an oven at 103±2 °C for 4 h The water content was based on weight differences before and after the drying of three replicates for each sample (ISO
6496, 1999) Salt content was measured with the Volhard Titrino method according to AOAC (2000).
Drip measurements and water holding capacity
Experiment I Drip was evaluated through the storage by measuring the weight of the fish before and after packaging The drip (%) was then calculated as the ratio of the weight of the water lost during storage to the original weight of the fish
Water holding capacity (WHC) of the minced fish was determined using a centrifugation method described by Eide et al (1982) Approximately 2 g of accurately weighed minced fish were centrifuged at 210 × g for 5 minutes at 0–5 °C using a Heraeus Biofuge Stratos centrifuge (Kendro Laboratory Products, USA).
At 15 °C, the weight loss observed during centrifugation (Δm_centrifuge) was treated as water loss, with no corrections for other components even in fish with high fat content Water-holding capacity (WHC) was calculated as the ratio of the water retained in the sample to the initial water mass before centrifugation, i.e., WHC = retained water / initial water mass.
Temperature measurements
Figure 4 Temperature in the climate chamber of the real temperature simulation (RTS)
Figure 4 illustrates the real air freight temperature simulation alongside the steady climate chamber conditions Boxes arrived at Matís facilities at 08:45 on February 25, 2009, and the RTS groups were placed in a climate chamber with an initial air temperature of 4.6 ± 3.0 °C After 12 hours, the temperature was reduced to -0.6 ± 0.7 °C for another 12 hours, then increased to 9.8 ± 2.2 °C for 6 hours, and finally held steady at -0.5 ± 0.5 °C for the remaining storage period The groups subjected to steady-temperature conditions were kept in a climate chamber at -1.2 ± 0.2 °C throughout the entire storage period.
Figures 5–8 show that the bottom corner of the EPS box is the most sensitive to temperature load, exhibiting the fastest response to temperature changes across all four cases This makes the corner region the critical hotspot for thermal effects in the studied enclosure.
Days from packaging top ‐ 422 centre ‐ 27 bottom ‐ 418 corner ‐ 425
Boxes inserted in climate chamber at 4.6°C for 12 hours
Climate chamber at 9.8°C for 6 hours
Climate chamber at ‐0.5°C for the remaining storage period
Figure 5 Group A (CBC-RTS-GP) – CBC cooled and packed with a gel pack
Days from packaging top ‐ 440 centre ‐ 444 bottom ‐ 439 corner ‐ 468
Boxes inserted in climate chamber at 4.6°C for 12 hours
Climate chamber at 9.8°C for 6 hours
Climate chamber at ‐0.5°C for the remaining storage period
Figure 6 Group B (CBC-RTS) – CBC cooled and packed without gel pack
Days from packaging top ‐ 378 centre ‐182 bottom ‐ 176 corner ‐ 165
Boxes inserted in climate chamber at 4.6°C for 12 hours
Climate chamber at 9.8°C for 6 hours
Climate chamber at ‐0.5°C for the remaining storage period
Figure 7 Group C (LC-RTS-GP) – liquid cooled and then packed with a gel pack
Days from packaging top ‐ 441 centre ‐483 bottom ‐ 479 corner ‐ 473
Boxes inserted in climate chamber at 4.6°C for 12 hours
Climate chamber at 9.8°C for 6 hours
Climate chamber at ‐0.5°C for the remaining storage period
Figure 8 Group D (NC-RTS-GP) – No cooling during processing and packed with a gel pack
Table 5 shows the average product temperatures, indicating that the group cooled with liquid cooling before packaging (C) maintained a lower temperature during the storage period than the group that received no cooling (D) Specifically, the average product temperature of the liquid-cooled group was 0.4 ± 0.6 °C, compared with 1.1 ± 0.5 °C for the untreated group The CBC cooling groups achieved even lower temperatures, at -0.1 ± 0.3 °C with a gel pack and 0.2 ± 0.5 °C without a gel pack.
Figures 5–8 confirm that CBC cooling yields a lower initial temperature in the treated products and enhances their thermal stability The CBC-treated items are more resistant to ambient temperature fluctuations and respond more slowly to changes in surrounding conditions.
During storage, the gel pack appears to modestly reduce the average product temperature, with the CBC-cooled group that included a gel pack showing slightly lower temperatures than the same group without one The gel pack may have a larger effect in groups C and D, where the product temperature at packaging is higher than in the CBC-cooled groups There is a clear difference in initial product temperature between groups that received CBC cooling and those that received only liquid cooling or no cooling, with packaging temperatures in groups C and D about 2–3 °C lower The top layer also showed lower temperatures at packaging and during storage compared with other locations, likely due to the gel pack being placed on top of the product inside the boxes Interestingly, temperatures in all groups reached a minimum after roughly 6–7 days and then began to rise, even though the ambient temperature was lower than the product temperature, probably due to exothermic reactions during spoilage.
Table 5 Average product temperature and standard deviation during storage group mean
Figures 9-10 show the product temperature of the groups which were kept at a steady ambient temperature of -1.2±0.2 °C during the entire storage period at Matís facilities
Days from packaging top ‐ 445 centre ‐ 432 bottom ‐ 460 corner ‐ 458
Product put in climate chamber
Mean ambient temperature ‐1.2°C for the remaining storage period.
Temperature logger at the corner shows rising temperature during the storage at transporter's chilled store
Figure 9 Group E (CBC-S-GP) - CBC cooled and packed with a gel pack
Days from packaging top ‐ 4 centre ‐ 153 bottom ‐ 462 corner ‐426
Product put in climate chamber
Mean ambient temperature ‐1.2°C for the remaining storage period.
Figure 10 Group F (CBC-S) – CBC cooled and packed without gel pack
Similar to the pattern observed after day 7 in the RTS groups, the groups maintained at a steady temperature exhibit a rising product temperature despite a low and steady ambient temperature, which can be explained by exothermic reactions occurring during the spoilage process.
Figures 9–10 show the temperature distribution inside the package, with the bottom corner recording the lowest temperature and being most exposed to ambient conditions The top of the products is cooler than the center and the bottom of the box, a pattern likely due to the gel pack located on top of the product These findings illustrate how gel pack placement and package geometry influence thermal performance during storage and transport.
Comparison of temperature profiles among groups
CBC‐RTS‐GP CBC‐RTS LC‐RTS‐GP NC‐RTS‐GP CBC‐S‐GP CBC‐S
Climate chamber at 4.6±3.0°C for 12 hours Climate chamber at 9.8±2.2°C for 6 hours
Figure 11 Average product temperature history of all the groups during transportation and storage at Matís facilities
Figure 11 illustrates the product temperature history for two groups: those subjected to real temperature simulation (RTS) and those stored at a constant subzero temperature (-1.2 ± 0.2 °C) during storage at Matís facilities.
The influence of CBC cooling can clearly be seen on Figure 11 as the group CBC-RTS-
GP shows significantly lower temperature profile during the temperature load applied for the first two days of the experiment
The gel pack helps dampen the temperature response of products under a temperature load, as shown by the comparison of CBC-RTS-GP and CBC-RTS When comparing the same groups, the gel pack maintains a lower storage temperature throughout the storage period, with end-of-storage temperatures of 0.06 °C for CBC-RTS-GP versus 0.44 °C for CBC-RTS Liquid cooling similarly limits temperature rise during storage, keeping a lower temperature throughout than no cooling, even though the packaging temperature at the start was similar to the no-cooling condition.
Sensory evaluation
Figure 12 shows how the samples were characterized by sensory attributes In total, 95% of the sensory variation was explained by the first two principal components, with the main variation among samples arising from storage time Early in storage, cod exhibited attributes such as sweet and metallic flavors and sweet and shellfish odors, located on the right in the upper part of Figure 1b for samples after 1–2 days (Figure 12a) As storage time progressed, these attributes waned while vanilla odor and juicy texture became more characteristic; later, potato odor and white precipitation appeared in the lower part (Figure 12b) At the end of storage, cod showed attributes like TMA (trimethylamine) and sour flavors and odors, located on the left in the upper region and used to describe end-of-storage samples The NC-RTS-GP group retained freshness characteristics longer, being comparable to other groups after six days of storage as shown in Figure 12a In contrast, LC-RTS-GP lost freshness characteristics faster and exhibited more spoilage attributes after eight days (LC-RTS-GP results after 10 days were removed from the multivariate analysis due to extreme spoilage values).
Tables A-D in appendix show in more detail how the sample groups were characterized by sensory attributes
• CBC-RTS-GP-d10 CBC-RTS-GP-d01 •
• CBC-RTS-GP-d06 CBC-RTS-GP-d08 •
• NC-RTS-GP-d06 NC-RTS-GP-d08 •
X-Loadings f-off • o-sweet • a-discoloured • f-sweet • f-metallic •
• o-vanilla o-potatoes • a-precipitation • o-cloth • • o-sour
Figure 12 presents a principal component analysis (PCA) of sensory quality attributes—odour (o-), appearance (a-), flavour (f-), and texture (t-)—for sample groups across storage time The PC1–PC2 score plot captures the majority of variation, with 90% explained by PC1 and 5% by PC2, highlighting how storage duration and cooling strategy influence sensory outcomes The panel includes a) scores and b) X-loadings, illustrating which attributes drive separation among samples Cooling conditions assessed are Combined blast and contact cooling (CBC), Real temperature simulation (RTS), Liquid cooling (LC), No cooling (NC), Gel pack (GP), with storage at −1 °C (S).
Figure 13 shows how the Torry freshness score changes with storage time A Torry score around 7 indicates the fish has lost most of its freshness odour and flavour characteristics and has a rather neutral odour and flavour (Shewan et al 1953) The limits were obtained after six days for LC-RTS-GP, after eight days for CBC-S, and after around seven days for the other sample groups When the average Torry score is around 5.5, most sensory panellists detect spoilage attributes, and these limits have been used as the limits for consumption at Matis (see Olafsdottir et al 2006) Accordingly, the maximum shelf life was 8–9 days for LC-RTS-GP, 9–10 days for CBC-RTS-GP and CBC-RTS, and 12 days for CBC-S-GP and CBC-S, with NC-RTS-GP approaching these limits on day 10.
Figure 14 demonstrates how the fish's sweet flavor diminishes with storage time; a sweetness score of about 25–30 indicates that most of the characteristic sweet flavor has been lost The time to reach these limits varies by treatment: LC-RTS-GP around seven days, CBC-S about nine days, and roughly seven to nine days for the other groups, which is about one day longer than what the Torry score indicated.
Figure 13 Average Torry freshness scores
(CBC: Combined blast and contact cooling,
RTS: Real temperature simulation LC: Liquid cooling, NC: No cooling, GP: Gel pack, S:
Figure 14 Average QDA scores of sweet flavour (CBC: Combined blast and contact cooling, RTS: Real temperature simulation LC: Liquid cooling, NC: No cooling, GP: Gel pack, S: Storage at -1 °C)
Figures 15–20 show how odour and flavour attributes related to spoilage change with storage time End of shelf life is usually determined when sensory attributes associated with spoilage become evident When the average QDA score for those attributes exceeds 20 on a 0–100 scale, most panellists detect them (Bonilla et al 2005; Magnússon et al 2006) According to this criterion, the maximum shelf life was 8–9 days for LC-RTS-GP, nine days for CBC-RTS, ten days for CBC-RTS-GP, and around 12 days for CBC-S-GP and CBC-S.
13 days, but NC-RTS-GP was approaching end of shelf life after 10 days These results are in agreement with the results from the Torry scores
Figure 15 Average QDA scores of table cloth odour (CBC: Combined blast and contact cooling, RTS: Real temperature simulation LC:
Liquid cooling, NC: No cooling, GP: Gel pack,
Figure 16 Average QDA scores of TMA odour (CBC: Combined blast and contact cooling, RTS: Real temperature simulation LC: Liquid cooling, NC: No cooling, GP: Gel pack, S: Storage at -1 °C)
Figure 17 Average QDA scores of sour odour
(CBC: Combined blast and contact cooling,
RTS: Real temperature simulation LC: Liquid cooling, NC: No cooling, GP: Gel pack, S:
Figure 18 Average QDA scores of sour flavor (CBC: Combined blast and contact cooling, RTS: Real temperature simulation LC: Liquid cooling, NC: No cooling, GP: Gel pack, S: Storage at -1 °C)
Figure 19 Average QDA scores of TMA flavor
(CBC: Combined blast and contact cooling,
RTS: Real temperature simulation LC: Liquid cooling, NC: No cooling, GP: Gel pack, S:
Figure 20 Average QDA scores of off- flavor (CBC: Combined blast and contact cooling, RTS: Real temperature simulation LC: Liquid cooling, NC: No cooling, GP: Gel pack, S: Storage at -1 °C)
Table 6 compares the freshness period with the maximum shelf life of fish The freshness period ends when the fish has lost its freshness characteristics and reached the neutral phase, while the maximum shelf life ends when odour and flavour attributes related to spoilage become evident.
Table 6 summarizes the freshness period (in days) and the maximum shelf life (in days) as determined by sensory evaluation for six cooling and storage treatments: CBC (Combined blast and contact cooling), RTS (Real temperature simulation), LC (Liquid cooling), NC (No cooling), GP (Gel pack), and S (Storage at -1 °C) The results show how each treatment influences product aging as judged by sensory criteria, enabling a direct comparison of effectiveness in preserving quality during storage This breakdown helps identify the most effective cooling strategy for extending both freshness and shelf life.
Treatments applied to the groups did not alter the sensory characteristics of the samples beyond affecting the length of the freshness period and the maximum shelf life, as estimated from freshness- and spoilage-related odour and flavour attributes Specifically, brine immersion shortened the freshness period and maximum shelf life by two days compared with untreated loins (LC-RTS-GP vs NC-RTS-GP), likely due to brine contaminated by spoilage bacteria CBC cooling extended the freshness period by two days and the shelf life by one day relative to loins receiving the same treatment except CBC cooling (CBC-RTS-GP vs LC-RTS-GP) However, due to the brine immersion, CBC cooling did not prolong shelf life compared with loins not receiving this treatment The shelf life of untreated loins also exceeded that of the CBC groups The use of a gel pack had no significant effect on freshness period or shelf life (CBC-RTS-GP vs CBC-RTS; CBC-S-GP vs CBC-S) Storage at -1 °C (CBC-S-GP and CBC-S) prolonged shelf life by about three days compared with storage under simulated air freight temperature, with only a tendency for a longer freshness period observed.
Microbial measurements
Results from microbial counts (Figures 21–25) show that cod processed two days post-catch had low microbial counts in the control group on the day of processing (d0) for newly filleted cod fillets Treatment with liquid cooling and CBC led to higher microbial counts compared with the control The brine used was heavily contaminated with bacteria, notably Photobacterium phosphoreum, but bacterial numbers in the brine did not increase during the working day, apart from a slight uptick in P phosphoreum.
Microbial counts at the start of storage were lower in the no cooling group (NC-RTS-GP) than in the liquid cooling group (LC-RTS-GP) Thereafter, there were only minor differences between groups in total viable counts (TVC), H2S-producing bacteria, and P phosphoreum Pseudomonads counts were lowest up to day 6 in both the no cooling and liquid cooling groups, while during this period they were higher in the liquid cooling group than in the no cooling group These findings strongly suggest that fillets became contaminated from the brine used for liquid cooling, which carried high microbial loads (Figure 21).
Brine CBC-GP LC-GP Control-GP
Lo g 10 ba ct er ia /g
Figure 21 Microbial counts of cod loins and brine (CBC: Combined blast and contact cooling, LC: Liquid cooling, GP: Gel pack)
Microbial counts were generally lower in CBC-RTS samples when a gel pack was used compared with those without a gel pack, with the most noticeable reductions seen for H2S-producing bacteria and P phosphoreum during storage By contrast, in the CBC-S groups there was no marked difference in microbial counts whether a gel pack was used or not.
Lower microbial counts were generally observed in the CBC-S group compared with the CBC-RTS group, with the difference most notable for P phosphoreum The CBC-S group also experienced a lower storage temperature, with the initial temperature close to -1 °C (Figure 11).
CBC-RTS-GP CBC-RTS LC-RTS-GP NC-RTS-GP TVC
CBC-RTS-GP CBC-RTS CBC-S-GP CBC-S TVC
Figure 22 shows the total viable counts (TVC) in cod loins, with average values derived from triplicate samples and error bars indicating the standard deviation The figure compares several cooling and storage methods: CBC (Combined blast and contact cooling), RTS (Real temperature simulation), LC (Liquid cooling), NC (No cooling), GP (Gel pack), and S (Storage at -1 °C).
CBC-RTS-GP CBC-RTS LC-RTS-GP NC-RTS-GP
CBC-RTS-GP CBC-RTS CBC-S-GP CBC-S
Figure 23 illustrates the growth of hydrogen sulfide (H2S)-producing bacteria in cod loins, with the results shown as the average of triplicate samples and error bars representing standard deviation The figure compares several cooling strategies, including combined blast and contact cooling (CBC), real temperature simulation (RTS), liquid cooling (LC), no cooling (NC), gel pack (GP), and storage at -1 °C (S).
CBC-RTS-GP CBC-RTS LC-RTS-GP NC-RTS-GP Pp
CBC-RTS-GP CBC-RTS CBC-S-GP CBC-S Pp
Figure 24 shows the growth of Photobacterium phosphoreum in cod loins, with mean values from triplicate samples and error bars representing standard deviation The figure compares several cooling and storage strategies—CBC (Combined Blast and Contact Cooling), RTS (Real Temperature Simulation), LC (Liquid Cooling), NC (No Cooling), GP (Gel Pack), and S (Storage at −1 °C)—to illustrate how each method affects bacterial growth during storage.
CBC-RTS-GP CBC-RTS LC-RTS-GP NC-RTS-GP Pseudomonads
CBC-RTS-GP CBC-RTS CBC-S-GP CBC-S Pseudomonads
Figure 25 shows the growth of presumptive pseudomonads in cod loins, with average values from triplicate samples and error bars representing standard deviation The legend defines the cooling treatments as CBC (combined blast and contact cooling), RTS (real temperature simulation), LC (liquid cooling), NC (no cooling), GP (gel pack), and S (storage at -1 °C).
Chemical measurements
Total Volatile Base Nitrogen (TVB-N) and Trimethylamine (TMA)
Results from TVB-N and TMA measurements (Figures 26–27) are consistent with microbial counts, confirming parallel spoilage trends The NC-RTS-GP group, which received no cooling, showed the lowest TVB-N and TMA values, whereas the LC-RTS-GP group had considerably higher values Implementing gel packs slowed TVB-N and TMA formation, as evidenced by the contrast between CBC-RTS-GP and CBC-RTS.
The CBC-S group exhibited lower values than the CBC-RTS group, which is linked to the lower storage temperature applied to CBC-S The initial temperature for CBC-S was close to -1 °C (Figure 11).
CBC-RTS-GP CBC-RTS CBC-S-GP CBC-S
Figure 26 shows Total Volatile Base Nitrogen (TVB-N) levels in cod loins, with values representing the average of triplicate samples and error bars indicating standard deviation; the figure compares several cooling and storage treatments, including CBC (Combined blast and contact cooling), RTS (Real temperature simulation), LC (Liquid cooling), NC (No cooling), GP (Gel pack), and S (Storage at -1 °C).
CBC-RTS-GP CBC-RTS CBC-S-GP CBC-S
Figure 27 shows trimethylamine (TMA) in cod loins, with average values from triplicate samples and error bars representing the standard deviation The data compare several cooling and storage conditions, including CBC (combined blast and contact cooling), RTS (real temperature simulation), LC (liquid cooling), NC (no cooling), GP (gel pack), and S (storage at -1 °C) pH measurements were recorded alongside the TMA assessment.
Overall, pH tended to rise with storage time (Figure 28) Cooling method and storage conditions generally did not have significant effects, except on day 13, when pH was higher in CBC-cooled loins stored under temperature fluctuations without a gel pack.
CBC-RTS-GP CBC-RTS CBC-S-GP CBC-S
Figure 28 Acidity (pH) in cod loins Average values of triplicate samples are shown Error bars show
SD (CBC: Combined blast and contact cooling, RTS: Real temperature simulation LC: Liquid cooling, NC: No cooling, GP: Gel pack, S: Storage at -1°C)
Immersing the loins in a liquid containing 2.2% NaCl slightly increased their salt content, with immersed loins displaying 0.3–0.4% NaCl compared to 0.2% in the reference group (NC-RTS-GP); this difference is shown in Figure 29 Throughout storage, water content remained similar across all groups, ranging from 81% to 82% (Figure 30).
Figure 29 Salt content of cod loins that were immersed in liquid containing salt during processing Reference loins were packed without prior cooling (C: No cooling, LC: Liquid cooling, CBC: Combined blast and contact cooling, GP: Gel pack, RTS: Real temperature simulation, S: Steady temperature)
Figure 30 Changes in water content of cod loins that treated by different cooling and storage conditions Reference loins were packed without prior cooling (C: No cooling, LC: Liquid cooling, CBC: Combined blast and contact cooling, GP: Gel pack, RTS: Real temperature simulation, S: Steady temperature).
Drip and water holding capacity (WHC)
Drip loss from cod loins increased with storage time across all groups, rising from approximately 1% to 3–4% Notably, loins immersed only before packaging (LC-GP-RTS) exhibited about 3% drip loss on day 1 and continued to rise to roughly 4% during storage (Figure 31).
Figure 31 shows the changes in drip loss for cod loins subjected to different cooling and storage conditions, with reference loins packed without prior cooling The cooling conditions are labeled as C: No cooling, LC: Liquid cooling, CBC: Combined blast and contact cooling, GP: Gel pack, RTS: Real temperature simulation, and S: Steady temperature Black columns indicate that the drip may be underestimated because the loins were slightly frozen at the time of sampling.
During storage, the muscle's water-holding capacity (WHC) tended to decrease as muscle degradation increased (Figure 32) By the end of the storage period, WHC showed a slight rise These changes are attributed to increased drip loss; as more loosely bound water leaked from the muscle, the fraction of tightly bound water rose, producing the observed WHC decline followed by a modest rebound.
Figure 32 documents changes in the water holding capacity (WHC) of cod loins subjected to different cooling and storage conditions, with reference loins packed without prior cooling The treatments evaluated are no cooling (C), liquid cooling (LC), combined blast and contact cooling (CBC), gel pack (GP), real temperature simulation (RTS), and steady temperature (S) Black columns indicate that drip loss may be underestimated because the loins were apparently slightly frozen during sampling.
4 RESULTS AND DISCUSSION: EXPERIMENT II
Temperature control during air and sea transport from Iceland to Germany
During processing, fish fillets were chilled in CBC equipment to -0.9 to -0.5 °C Temperature mapping was conducted in eight EPS boxes, with cooling gel packs placed on top of the loins in four of them Half the boxes were exported by air freight and the other half by sea, and temperature fluctuations during transportation were analyzed to identify which segments of the transport chain are most associated with product quality loss Product and ambient temperatures were monitored, resulting in temperature maps for two pallets, one transferred by sea and the other by air freight.
Four Onset TidBit temperature loggers were deployed around the air‑freight pallet to monitor temperature at multiple positions—on the pallet surface, at the top centre, at the top corner, at the bottom corner, and between boxes at approximately 0.5 m height Four DS1922L iButton temperature data loggers were placed inside each of the eight EPS boxes (Figure 33) In addition, two iButton temperature data loggers were mounted on the outer surfaces of the EPS boxes.
Figure 33 DS1922L ibutton logger at the bottom centre of an EPS box
iButton temperature data loggers placed inside the EPS boxes were located at the bottom corner, bottom centre, mid centre and upper centre (below the top loin); the three loggers on the outer surface were positioned as Figure 34 shows The EPS boxes are designed for 5 kg of fresh fish products.
Figure 34 EPS boxes, Onset Tidbit and DS1922L ibutton temperature data loggers on outer surface
Results of the temperature mapping from packaging in Iceland to arrival in Bremerhaven by air freight transportation are shown in Figures 35 - 38
Reason for this: logger located in between boxes on pallet
Figure 35 presents ambient air temperature on the pallet, recorded by four Onset UTBI-001 temperature loggers placed on the pallet surface at the top center, top corner, bottom corner, and between boxes, alongside a HOBO U12 humidity and temperature logger FG1-2230536 denotes the temperature measurement from the HOBO logger, while FG1-RH_2230536 denotes the relative humidity measurement from the same device.
Figure 35 highlights hazardous parts of the air freight chain in terms of ambient temperature: on the Dalvík–Reykjavík leg, ambient temperatures were below 0 °C; from Reykjavík onward to Bremerhaven, temperatures were most of the time above 5 °C; and the most pronounced temperature fluctuations, including the highest ambient temperature, occurred on the Reykjavík–Cologne leg.
9:30 am (CET) Arrival in Bremerhaven
11:30 am Delivery of product to Grandi
9:25 pm (CET) Arrival in Cologne
In storage over night at Flytjandi in Reykjavík
Delivery to HB Grandi, Reykjavik 11:30 AM
At Matis - more loggers applied
Delivery to Bluebird Cargo, Keflavík 2:30 PM
Arrival in Cologne 9:25 PM (CET)
Arrival in Bremerhaven 9:30 AM (CET)
Figure 36 illustrates the temperature inside the EPS boxes and on their exterior surfaces during land and air transport Four DS1922L iButton temperature data loggers were placed inside each EPS box, for a total of sixteen loggers across four boxes, while two loggers were attached to the exterior surfaces of the boxes Eight loggers monitored ambient air temperature, and the average ambient temperature was calculated from these eight readings.
Figure 36 illustrates the temperature profile inside and outside the boxes during air freight transport, showing how the product temperature rises in parallel with high ambient temperatures The data indicate a direct relationship between external conditions and the shipment’s internal heat exposure, with temperature beginning to rise as aircraft loading starts and continuing to increase throughout the flight The rise persists until landing in Cologne, highlighting how ambient temperature and loading timing influence the thermal exposure of air-freight shipments.
Figure 37 shows the temperature inside the boxes during sea transport, indicating relatively steady conditions throughout the journey The ambient air temperature was below 0 °C for most of the trip, but as external temperatures rose, the product temperature increased accordingly A temperature rise was first observed upon arrival in Esbjerg.
9:00 am (CET)Arrival in Esbjerg
11:45 am (CET)Arrival in Bremerhaven
Figure 37 documents the temperature inside and on the exterior surfaces of EPS boxes during land and sea transport Four DS1922L iButton temperature data loggers were placed inside each EPS box, totaling sixteen loggers across four boxes, while two loggers were positioned on the exterior surfaces of the boxes The average ambient temperature was calculated from eight ambient temperature loggers.
Figure 38 illustrates the temperature evolution for air freight and sea transport The data show that temperature rose earlier in air freight, causing the product temperature to begin fluctuating after about 42% of the total transit time and to exceed zero degrees after roughly 64% In contrast, during sea transport the product temperature began rising after about 66% of the journey and reached 0 °C after about 81% of the time At the final destination, the average product temperature was 3 degrees lower for sea transport than for air freight.
Results show that there is hardly any difference in temperature between boxes containing a gel pack (groups FG and FS) and boxes without a gel pack (groups GG and GS) The negligible temperature difference suggests that the gel pack did not significantly influence temperature under the tested conditions.
Percent of total transportation time
Figure 38 shows the average temperature inside EPS boxes under two conditions: with a gel pack on top of the fillets (FS and FG) and without a gel pack (GS and GG) A total of four data loggers were placed inside each box to monitor temperature, enabling a direct comparison of thermal performance across the four groups.
The main results from this experiment show that temperature fluctuations were larger and more frequent for products transported by air than by sea, and ambient air temperature fluctuations led to increased product temperatures, with air freight shipped cod loins arriving in Bremerhaven exceeding 4 °C.
By comparing product temperature at the top, centre and bottom inside boxes, air freight shows temperatures above 0 °C for 35% of the transport time, with peaks reaching 4 °C, while refrigerated sea-container transport kept temperatures above 0 °C for 18% of the journey and never exceeded 1 °C.
On arrival at TTZ in Bremerhaven, the temperature of fish transported by air freight remained relatively high (Table 7) For some fillets, temperatures surpassed 4 °C, which is the rejection point for most fish retailers Gel packs did not provide a significant cooling effect in this experiment, possibly due to the small size of the gel packs.
(150 g) and secondly that the fish fillets were well pre-chilled before packaging (-0.9 to -
0.5 °C) The temperature of sea transported fish was significantly lower at arrival at TTZ
(Table 8) Surface temperature in boxes with a gel pack was slightly lower than that in boxes without a gel pack
Table 7 Temperatures (°C) at arrival (26th Feb 2009, 9:30 CET, air freight) P1 to P3 represent three different measuring positions in each box
FG 1 (with gel pack) Surface 4.2 2.3 3.5 3.3
GG 1 (without gel pack) Surface 4.1 4.2 2.5 3.6
Table 8 Temperatures (°C) at arrival (2nd March 2009, 11:45 CET, sea freight) P1 to P3 represent three different measuring positions in each box
FS 1 (with gel pack) Surface 1.2 0.9 0.6 0.9
FS 2 (with gel pack) Surface 2.0 1.6 1.5 1.7
GS 1 (without gel pack) Surface 2.9 2.7 2.7 2.8
GS9 (without gel pack) Surface 2.2 1.9 1.6 1.9
Microbial and chemical analysis
Microbial growth in cod loins transported by air freight and retail-packed under air or
MA 2 days post process is shown in Figure 39 MA-packaging generally delayed the product microbiota, especially pseudomonads and H2S-producing bacteria
Photobacterium phosphoreum (Pp) growth, evaluated by a quantitative PCR method in few samples obtained from Bremerhaven, showed a similar behaviour between air- and
MA-packed products, reaching log 7/g about 11 days after retail-packaging
Pseudomonad counts in air-stored products were similarly high by day 11, while H2S-producing bacteria reached that level three days later Based on the microbiological rejection criterion of log 7/g for spoilage bacteria, the shelf life of these products was probably reached on or near day 11 Generally, the initial use of the gel pack in EPS boxes (FG groups) during export had little influence, with most FG and GG curves evolving together.
FG-air GG-air FG-MAP GG-MAP
Figure 39 Microbial growth in cod loins transported by air freight and retail-packed under air or
MA, two days post-process, measurements quantified the total viable psychrotrophic counts (TVC) and the counts of Photobacterium phosphoreum (Pp), H2S-producing bacteria, and pseudomonads The values shown are the mean of duplicate samples, and the error bars represent standard deviation (SD).
During sea shipments, cod products experienced limited microbial growth for most of the journey due to consistently low temperatures (Figure 40) A rise in Pp on repackaging day (d0) compared with pseudomonads and H2S‑producing bacteria indicated that late-journey temperature increases particularly favored Pp After transport, retail packaging and subsequent air storage at slightly higher temperatures promoted rapid growth of spoilage bacteria, pushing populations toward critical levels.
7/g for pseudomonads and H2S-producing bacteria) after 8 days The results show that P phosphoreum had reached this level on day 10, again with a similar growth in all groups
However, more sampling points at earlier storage would have given a clearer picture of
Pp behaviour Proliferation of pseudomonads and H2S-producing bacteria in MA-packed products was delayed resulting in a probably longer shelf life (10-15 days) Pp levels were close to log 7/g on day 15
FS-air GS-air FS-MAP GS-MAP
Figure 40 Microbial growth in cod loins transported by sea freight and retail-packed under air or
Six days after processing under modified-atmosphere packaging (MA), total viable psychrotrophic counts (TVC) and counts of Photobacterium phosphoreum (Pp), hydrogen sulfide-producing bacteria, and pseudomonads were recorded The results are the mean values of duplicate samples, with error bars indicating the standard deviation.
Comparing sea freight and air freight for transporting fish products shows that the steady, low-temperature profile of sea freight, in contrast to the fluctuating temperatures typical of air freight, tends to preserve higher quality at the delivery site despite longer transit times This can be explained by the physiology of spoilage bacteria: in air-freighted fish, bacteria reach exponential growth around six days after processing (four days after repackaging), whereas sea-freighted fish remain in a late lag phase, slowing spoilage and maintaining quality.
At the same processing phase, cod loins were assessed to compare sea freight versus air freight performance Based on microbial counts, the estimated shelf life of cod loins shipped by air freight was about 13 days, with sea freight showing distinct microbial dynamics that could alter the overall shelf life under the studied conditions These results highlight the impact of transport mode and timing on freshness, safety, and product quality for cod loins in international shipping.
Repackaged fish stored under air conditions had a shelf life of 14 days, while modified-atmosphere (MA) stored fish lasted 16–21 days after sea-transported repackaging TVB-N content was measured at only a few sampling points and rose to higher levels at the end of storage (Figure 41) TVB-N production progressed slightly faster in air-stored fish than MA-stored fish when repackaged after air freight, but there was no difference between air- and MA-stored fish after sea freight.
Flesh pH changes were monitored more regularly (Figure 41 and Figure 42) A similar pH rise was observed in air-packaged fish regardless of transport mode, with the pH reaching about 7.1 after 13 days for air freight and 14 days for sea freight post-process.
MAP reduces the initial pH of cod loins by about 0.2 pH units, then the pH rises as amine degradation compounds accumulate Generally, sensory rejection of MAP-packaged cod is associated with a lower pH of about 6.8–6.9, compared with 7.0–7.1 in acceptable products, due to the buffering effect of CO2 dissolving into the muscle water phase.
FG-air GG-air FG-MAP GG-MAP
FS-air GS-air FS-MAP GS-MAP
Figure 41 compares TVB-N content in cod loins after transport by air (left) versus sea freight (right), with retail packaging in either ambient air or modified atmosphere (MA) and measurements taken at 2 and 6 days post-processing The values shown are the averages of duplicate samples, highlighting how transport mode, packaging atmosphere, and storage duration influence TVB-N levels in the cod loins.
FG-air GG-air FG-MAP GG-MAP
FS-air GS-air FS-MAP GS-MAP
Figure 42 compares pH development in cod loins during transport by air (left panel) and sea freight (right panel), as well as in retail-packaged products stored under air or modified atmosphere (MA) after 2 or 6 days post-processing, with the average pH values shown from duplicate samples.
TVB-N content of 40–50 mg N/100 g at sensory rejection for CBC loins stored under superchilled conditions, versus 30 mg N/100 g in abused CBC loins (Figure 26), indicates that the shelf life of these products may be slightly shorter This effect is especially evident for fish transported by air freight and stored under air, which re-estimates the overall shelf life to 8 days for air-stored fish and 11 days for MA-stored fish when comparing TVB-N levels in air- and MA-stored fish Table 9 summarizes and compares the shelf-life estimates derived from the measured parameters. -**Support Pollinations.AI:** -🌸 **Ad** 🌸Powered by Pollinations.AI free text APIs [Support our mission](https://pollinations.ai/redirect/kofi) to keep AI accessible for everyone.
Table 9 presents the estimated shelf life and overall shelf life (in days) of retail-packed cod loins, showing how the transportation mode (air or sea freight prior to repackaging) and the atmospheric condition influence the product’s longevity.
Shelf life determination criteria Air freight Sea freight
TVB-N: 30-50 mg N/100 g 6 (8) 9 (11) 5-6 (11-12) 5-6 (11-12) pH: 7.0-7.1 (air), 6.8-6.9 (MAP) 9-10 (11-12) 11-14 (13-16) 7 (13) 9 (15) a overall shelf life from process time, including transportation time, given in parenthesis
Cod loins exhibit similar TVB-N values at arrival at TTZ, whether transported by air or by sea This observation suggests that extended sea freight times do not automatically compromise fish quality if proper temperature control is maintained.
45 maintained In fact, fish transported by sea showed the slowest quality deterioration according to TVB-N values
CBC cooling resulted in a significantly lower temperature profile during the temperature load applied for the first two days of the experiment The use of a gel pack lowered somewhat the temperature increase in the products when the temperature load was applied and lower temperature was maintained during the entire storage period with a gel pack As compared to no cooling, the use of liquid cooling contributed to maintaining a lower temperature during the entire storage period, even though the temperatures at packaging were similar