The effect of drying, pressure and processing time on the quality of liquid-smoked trout (Salmo gairdnerii) fillets

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1 Journal of the Science of Food and Agriculture J Sci Food Agric 85: (2005) DOI: /jsfa.2220 The effect of drying, pressure and processing time on the quality of liquid-smoked trout (Salmo gairdnerii) fillets Ilias Siskos, 1 Anastasios Zotos 1 and KD Anthony Taylor 2 1 Technological Educational Institution of Thessaloniki, School of Food Technology and Nutrition, Department of Food Technology, Thessaloniki, PO Box 141, Greece 2 Food Research Centre, Department of Biological Sciences, Faculty of Health and Life Sciences, University of Lincoln, UK Abstract: A new fish smoking process was applied using a combination of liquid smoke and steaming at pressures up to 1 bar above atmospheric. Processing yield, sensory analysis, instrumental colour measurement, available lysine and polynuclear aromatic hydrocarbons (PAHs) were estimated. The losses due to processing were quite reasonable (25 ± 4.9% to ± 3.9%) and slightly influenced by the process. The fillets processed at 2 bar steam pressure, for 30, 45 or 60 min and previously dried were assessed as highly acceptable regarding their firmness, colour, flavour and acceptability by panellists. The destruction of available lysine was not very high (21.1 ± 8.4%) and it was dependent upon the process. Depending on the method used, very low ( ng g 1 ) amounts of PAHs were found and were also dependent upon the process Society of Chemical Industry Keywords: smoked trout; liquid smoking; pressure; PAHs; lysine INTRODUCTION Smoking is one of the most ancient ways of preservation. No other way of fish processing can produce such characteristic colour and flavour. Traditionally, smoking of fish is usually conducted using either cold smoking at temperatures not exceeding 30 C or hot smoking in conditions causing thermal denaturation of proteins. 1 Smoking in northern European countries is conducted in several ways, which results in different characteristics of the final product. For example in Norway, salmon is salted for 3 to 4 days, air-dried for 10 to 12 h and finally cold smoked for 4 to 10 h, whilst the hot smoked trout are processed for 2 1 to 3 h at 2 80 C. 2 Liquid smoke is generally used as a flavouring agent. The following steps of processing are reported: brining at 3.3 C for 2 days in a fish:brine ratio of 1:7. 3 In some of the above experiments liquid smoke was mixed with brine in a concentration of 7 or 22 ml l 1. Alternatively, liquid smoke was applied by dipping brined fish for 1 min in either 100 or 50% aqueous liquid smoke solution. Fish fillets were dipped for 1 min in either 100, 50, 25 or 10% liquid smoke solution. The fish fillets were allowed to drain on stainless steel mesh screens before heat processing. 4 However, it has been reported that the use of liquid smoking does not provide products acceptable in terms of in colour, flavour and appearance. 5 One of the major problems with traditional smoking is the occurrence of polycyclic aromatic hydrocarbons, which are well-established as carcinogens. 6 It has been reported that a total concentration of polyaromatic hydrocarbons ranged between 200 and 480 µgkg 1 in liquid smoked products and 650 and 1200 µgkg 1 in traditionally cold smoked fillets. 7 A concentration of 1.97 µg benzo(a)pyrene [B(a)P] g 1 and 0.3 µg B(a)P g 1 flesh was detected in traditional smoked salmon-trout and smoked trout, respectively, and several other polycyclic aromatic compounds in the different ways of smoking tested. 8,9 A concentration of ng B(a)P g 1 was also found in smoked trout as well as other potential carcinogenic compounds. 10 It is well known that lysine participates with the ε-amino group in the early steps of Maillard reaction pathways. This reaction may occur at ambient or lower temperatures over time, although the rate of the reactions increases with temperature. Carbonyl compounds are present in fish in very low levels; however, they are present in smoke or arising from lipid oxidation. Thus, these carbonyls may react with the ε-nh 2 group of lysine in a similar manner to reducing sugars. The impact of these reactions is a reduction Correspondence to: Anastasios Zotos, Technological Educational Institution of Thessaloniki, Department of Food Technology, Thessaloniki, PO Box 141, Greece zotos@food.teithe.gr (Received 27 February 2004; revised version received 15 November 2004; accepted 9 February 2005) Published online 6 June Society of Chemical Industry. J Sci Food Agric /2005/$

2 Factors affecting quality of liquid-smoked trout fillets in protein utilization mainly by the reduction of lysine through its ε-amino group. 11 Thus, the aim of this work was to produce a smoked product using a new sort process, based on liquid smoke and supported by the application of different minor pressures. The study also examines the effect of the different applied processes on the sensory characteristics of the products, such as texture, colour and flavour, as well as on the quality of the products with respect to hygiene and nutritional aspects. MATERIALS AND METHODS Smoking process Farmed trout (24 samples) (Salmo gairdnerii) was purchased from Ioannina, Greece. Immediately after arrival, fish samples were stored in ice at 0 1 C. The average weight of the samples was 900 g to 1 kg. About 20 h after harvesting they were eviscerated, the head removed, and brined in 20% sodium chloride solution for 2 h. The brine temperature was kept low (5 C) in order to minimize microbiological contamination. Consequently, all samples were filleted and half of them were placed in drying racks for 16 h (the drying time was chosen based on a preliminary experiment described below) while the other half was immediately processed. The temperature of all samples (dried and non dried) before processing ranged between 18 and 22 C. The smoking process was conducted using a 10-L electrical commercial steamer, electronically modified in the workshop to improve its precision and to which 40 ml of liquid smoke (Nefeloudis SA, Lavrion, Greece) condensate was added and diluted in 2 L of tap water (2%) with a final temperature of 20 ± 2 C. Thereafter, the smoking process took place by the production of smoky steam of the liquid smoke solution. Samples were placed at one layer with the flesh facing the smoke liquid and processed at four different processing times: 15, 30, 45 and 60 min. Processing time was measured from the moment the required pressure in the steamer was achieved (approximately min). The applied steam pressure conditions were: 1 (atmospheric pressure), 1.5 and 2 bar and the consequent temperatures were 100, 104 and 115 C, respectively. After each process the liquid smoke solution was discarded and a new solution was used. After cooling the samples at room temperature, under sanitary conditions, the skin was removed from the fillets. The front part (6 8 cm) from each fillet was removed carefully, packed in polypropylene bags, stored at 5 C for 3 days and used for sensory analysis. The remainder of the fillets were homogenized at low temperature in order to avoid possible oxidation, mixed thoroughly and stored in polyethylene bags at 5 C until further analysis (approximately for 1 week) (Fig 1). The combination of the two factors mentioned above resulted in 12 different processing levels. These, combined with the two drying levels (dried and nondried), gave rise to 24 non-identical ways of processing as shown in Table 1. Pre-smoking determination to optimise drying time Trout purchased from the same fish farm were dried at 20 C and 70% relative humidity, under six different drying times (2, 4, 6, 8, 16, and 24 h). After drying, all fillets were smoked at the higher pressure used in this work (2 bar) for 45 min in order to produce a more distinguishable appearance for the panellists samples. After processing the fillets were left to equilibrate at room temperature and were packed in polyethylene bags and stored at 5 C for 3 days prior to sensory analysis. The sensory evaluation was performed as described below using 10 experienced panellists. Each panellist tested four samples and was asked to evaluate the same variables described below. 20 h at 0 to 1 C eviscerated and head removed brined in 20% sodium chloride solution for 2 h at 5 C drying for 16h at 20 C, 70% RH smoking (steaming) at 1, 1.5 and 2 bar pressure filleting smoking (steaming) at 1, 1.5 and 2 bar pressure cooled at room temperature, packed stored at 5 C Figure 1. A presentation of the entire process. Table 1. Processing plan and coded samples 1 bar (atmospheric pressure) 1.5 bar pressure 2 bar pressure Time (min) Dried A1 A2 A3 A4 A5 A6 A7 A8 A9 A10 A11 A12 Non-dried B1 B2 B3 B4 B5 B6 B7 B8 B9 B10 B11 B12 J Sci Food Agric 85: (2005) 2055

3 I Siskos, A Zotos, KDA Taylor Weight loss All samples were weighed before and after all processing steps. After processing fillets were left to equilibrate at room temperature before weighting. Instrumental colour measurement Colour was not measured directly on the fish fillet surface, because it was porous and difficult to achieve repeatability. Thus samples (approximately 3 5 mm in depth) were homogenised, mixed thoroughly and homogenised again until a fine and homogenous paste was achieved. All sample preparation procedures were made by taking precautions to avoid excess moisture, air and to achieve homogeneity of colour on the surface of the final fish paste. Each sample (10 g) was placed in a small black container (3 cm diameter and 3 cm height), with the instrument s spectrally pure glass on the top and measured on the instrument s white tray, care being taken to cover all the available volume. The colour measurement was conducted using a colour difference measuring instrument ( Micro Color LMC colorimeter, Dr Lange GmbH, Berlin, Germany) which is a tristimulus colorimeter with the following optical structure. A Ulbricht globe, in conjunction with xenon flash lamp which generates lighting conditions approximating those of standard illuminant D 65 and standard observed 10, served to diffusely illuminate the sample to be measured. The diffuse reflection of the sample at an angle of 8 was measured in accordance with German Industrial Standard DIN Each measurement was repeated five times for each sample. L, a and b values were measured and RI was calculated using the equation RI = a b The black container was cleaned and dried between the measurements of each sample. Available lysine analysis The available lysine was determined by the method described by Peterson and Warthesen. 13 Polycyclic aromatic hydrocarbons (PAHs) analysis Polycyclic aromatic hydrocarbons content was determined using two different methods, described by Larsson 14 and by Lawrence and Weber. 6 Both methods include digestion in methanolic solution of potassium hydroxide, partitioning in dimethyl sulfoxide or dimethylformamide and a column clean-up procedure. The chromatogram was obtained using a Focus GC (Thermofinnigan Italia SpA, Rodano, Milan, Italy) packed with a capillary 30-m column purchased from Alltech (Dearfield, IL, USA) with film thickness 0.25 µm and ID 0.32 mm. The method followed was: initial temperature 180 C for 3 min, rate 8 Cmin 1 until 320 C and 20 min isothermal. The injection and detection temperatures were 270 and 350 C, respectively. The carrier gas was helium with flow rate 3.5ml min 1. Sensory analysis Sensory evaluation was performed using 10 experienced panellist members of the academic staff for the different treatments. Therefore, the 10 panellists were used five times, testing five samples each time. Three sensory variables examined were: firmness, intensity of smoked colour and intensity of smoked flavour. Fifty students were also used to assess the overall acceptability of the different products. The intensity of all four variables was recorded on a 15-cm unstructured scale line. The line left end was marked at 0 cm for all variables and characterized as non firm, non-smoked flavour, white colour and extremely unacceptable, whereas the right end was marked at 15 cm as extremely firm, strong smoked flavour, brown and extremely acceptable. After the samples had been left for 30 min at room temperature, four segments of 5 cm diameter were placed in an odourless plastic container, covered with a watch glass and served to panellists. Samples were coded with three-digit random numbers and served in random order. Each panellist was served cooked samples after a 30-min break. Panellists evaluated samples in individual booths equipped with white lights. Statistical analysis Statistical analysis for the estimation of drying time was performed by the application of a balanced incomplete block design (BIBD) including t = 6treatments(6 different times), n = 10 replicates, b = panellists and k = 4 treatments per panellist. All different processed trout fillets were analysed using a two-factor ANOVA (one fixed treatments and one random panellists). A second balanced incomplete block design was applied on factors [dried/non-dried (2 levels), pressure conditions (3 levels), processing time (4 levels), Table 1)] including ultimately t = 25 combined treatments, n = 5 replicates per treatment, b = 50 panellists and k = 5 treatments per panellist (k < t). Thus, a combination of = 24 samples were achieved and the design was increased with a sample of smoked trout from the local market ( ) in order to conform to the particular plan obtained from Cochran and Cox. 15 Adjusted sensory mean scores were deduced for the 25 samples and, at this point, the 25th sample, obtained from the market, was excluded from further investigation. A three-way ANOVA for the three factors (Table 1) and their interaction terms was attempted on the set of sensory variables to detect the significant differences. Statistically significant differences between factor levels were tested using the Student Newman Keuls (SNK) test for comparison of level mean values. RESULTS AND DISCUSSION The chemical composition of the samples through out the whole study was relatively stable % moisture, % protein and % lipid, 2056 J Sci Food Agric 85: (2005)

4 Factors affecting quality of liquid-smoked trout fillets flavour firmness colour % loss bar pressure 1.5 bar pressure 2 bar pressure Drying time (h) min 30 min 45 min 60 min Processing time Figure 2. The effect of the different drying time prior to processing on flavour, firmness and colour of the processed trout fillets, after data standardization. indicating that small changes should probably be expected from farmed fish. Estimation of drying time The two-factor analysis of variance revealed significant differences for all variables considered (firmness, flavour, colour) between the samples that were dried at 20 C and at different times prior to processing. According to the SNK test the trout samples were separated into two groups: the first group comprised the samples that were dried for 2, 4, 6 and 8 h, while the second one the samples dried for 16 and 24 h prior to processing. No statistical differences was observed between the samples among each group; however, the samples of the second group were assessed by the panellists as more brown coloured and smoke flavoured than the samples of the first group (Fig 2). Therefore, the minimum drying time (16 h) prior to processing that led to the production of more acceptable products was chosen and used in this investigation for processing the dried fillets. Yield losses A 1.3 ± 1% gain in weight was observed due to brining in all trout samples. Statistical analysis revealed significant differences between the dried and nondried fillets prior to processing and between those that were processed under different steam pressure conditions. The mean yield loss of all samples was 21 ± 4.5% (mean loss ± standard deviation) throughout the entire process. The average yield loss of the fillets that were previously dried was ± 3.91%, which came from either the drying (9.6 ± 4.5%) or the smoking process (13.98 ± 3.32%). The non-dried fillets showed a 24 ± 4.04% average yield loss due to smoking. The lower losses observed on the dried fillets due to smoking were probably due to the formation of a protective dried surface that occurred during drying. The simultaneous increase in pressure and time increased yield losses. It was also observed Figure 3. Yield losses in line with time of processing and pressure conditions regardless of the drying conditions (error bars are standard deviations, n = 3). that losses in non-dried fillets showed an increasing trend due to increasing pressure conditions, while the average losses of dried fillets were stable at atmospheric and 1.5 bar steam pressure conditions (Fig 3). The mean yield due to processing in this work was relatively high (78.6%), despite the applied high temperatures, higher than those reported in the literature due to hot smoking. Thus, yields ranged from 68 to 53% due to hot smoking were reported in skin on (prevents losses), gutted without head fish produced in a traditional smoke house at lower temperatures. 16 This probably indicates that the sort processing time compensates well for the strong processing conditions. Analysis of sensory and colour results The most important factors influencing almost all the measured variables were the steam pressure conditions and the drying prior to smoking process. The processing time was also shown to have an effect on the properties of the final product but to a lesser extent. The significance of the main effects was tested through a three-way ANOVA in all measured variables. The drying factor was significant in all sensory variables (p < 5). The fillets that were previously dried showed stronger smoked flavour, were more intense brown colour, more firm and were more acceptable than the non-dried ones (Fig 4). The effect of moisture content on colour development is well known. Eichner and Karel 17 observed that an increase in water content resulted in a decrease in formation of coloured compounds. Yeo and Shibamoto 18 also reported the same effect. Steam pressure conditions also significantly influenced the sensory variables. All fillets that were processed at 2 bar steam pressure showed more intense smoked flavour and were more brown coloured. This was also statistically confirmed by the SNK test. This indicates that the extent of browning reactions was higher in the more intense process, as well as that the higher the applied pressure on the samples the J Sci Food Agric 85: (2005) 2057

5 I Siskos, A Zotos, KDA Taylor stronger the smoked flavour of the products (Fig 5). This should be expected since the higher the applied pressure the higher the temperature and consequently the higher the production of carbonyl and phenolic compounds that led either to more browning or to more flavoured final products. Smoked flavour and brown colour intensity were also influenced by the time of processing. Fillets processed for 45 and 60 min showed higher smoked flavour than those processed for 15 min, however, no significant differences in smoked flavour were observed between the samples processed either for 15 and 30 min or for 30, 45 and 60 min. Moreover, non-dried Drying process dried firmness flavour Colour Acceptability Figure 4. The effect of drying prior to processing on the sensory variables of the processed trout fillets, after data standardization. fillets processed for 60 min showed higher brown colour intensity than those processed for 15 min (Fig 6). Similar results were found by Fogliano et al, 19 working on a gluten glucose model system. They demonstrated that the production of colour increased with the time of processing. As expected, the acceptability of the products followed the above two variables. The trout fillets dried and then processed for 60 min at 2 bar steam pressure conditions were assessed as the most acceptable products by the panellists. All instrumental colour variables were affected by the steam pressure conditions. Lightness (L ) reduced according to SNK test due to increasing pressure (Table 2) and to drying process prior to processing. Thus, the L value of the non-dried products was (0.17) whilst that of the dried ones was (7) (p = 04). Similar results were reported by Morales and van Boekel 20 in several casein/sugar models where L values decreased significantly in the more severely heated samples indicating increased darkness. The L values found in this work were higher than those reported by other authors for smoked Atlantic salmon and this was probably due to homogenization before measuring. 21,22 All fillets processed at 2 bar steam pressure showed higher b values than those processed at 1 and 1.5 bar. The increase of pressure conditions seemed to produce fillets with a reverse trend in a values; however no significant differences were detected. These results indicate that increasing pressure conditions Flavour Colour flavour Colour Acceptability bar (atmospheric) pressure 1.5 bar steam pressure Pressure levels 2 bar steam pressure min 30 min 45 min 60 min Processing time Figure 5. The effect of the applied pressure on flavour and colour of the processed trout fillets, after data standardization. Figure 6. The effect of processing time on flavour, colour and acceptability of the processed trout fillets, after data standardization. Table 2. Effect of pressure conditions on a, b, L and RI values RI values L values a values b values 1 bar pressure 0.33 a (7) 69.4 b (1.44) 4.9 a (1.10) 14.8 a (0.69) 1.5 bar pressure 0.32 a (4) 67.7 a (1.48) 4.7 a (0.77) 14.7 ab (0.96) 2 bar pressure 0.24 b (6) 66.1 a (1.26) 3.8 a (2) 16.2 b (1.55) Values are means of fivefold determinations. Standard deviations are shown in parenthesis. Values marked with different superscript letters indicate statistical difference. p values were 04, 1 and 04 for RI, b and L values, respectively J Sci Food Agric 85: (2005)

6 Factors affecting quality of liquid-smoked trout fillets Table 3. Pooled means of available lysine content and losses due to processing Unprocessed 15 min 30 min 45 min 60 min 1 bar 1.5 bar 2 bar Protein (mg g 1 ) (0.25) (0.35) (0.40) (0.67) (0.83) (0.46) (0.61) (0.72) Loss (%) Data are means of triplicate determinations. Standard deviations are shown in parenthesis. led to more yellow and less red final products (Table 2). In addition, the redness index (RI) was also strongly correlated with smoked flavour and brown colour intensity, ( and respectively) indicating a negative relationship between the above sensory and instrumental measured variables. The above relationship can be interpreted as follows: as smoked flavour and brown colour increased due to more intense processing conditions, the redness index decreased leading to a less red and more yellow final products (Table 2). Available lysine content Statistically significant different results were obtained for available lysine due to drying prior to smoking. Thus, the pooled value of available lysine of the dried samples was (0.34) while that of the non-dried ones was (0.68) (p = 29). The average reduction in all samples was 21.1% ± 8.4. As shown in Table 3, the destruction of available lysine was shown as a possible trend due to all different parameters considered in the study. However, no statistical differences were identified between the samples. Shiau and Shue 23 investigated the effect of pre-frying times of canned tilapia meat on available lysine destruction and reported that the destruction increased with increasing time of processing. However, the destruction in this work was not as high as reported by Chen and Issenberg 24 who found 44.5% destruction due to smoking at 65 C for 10 h. Zotos et al 25 also reported 50 63% available lysine loss due to hot smoking of mackerel. PAHs The benzo(a)pyrene (BaP) concentration in the liquid smoke was 3.24 ng g 1 whilst, in the samples, the BaP content, as determined using the method described by Larsson, 14 ranged between 0.63 and 3.2ngg 1. The highest BaP content was found in the samples that were processed for 60 min and at 2 bar steam pressure conditions while the lowest was in the samples processed for 15 min at 1 bar (atmospheric pressure). This is in agreement with Chen and Lin 26 who observed that BaP content increased with increasing time of smoking of duck fillets. Relatively low BaP values were also found in the samples processed under 1.5 bar pressure (1.17 ng g 1 ). The levels of BaP content were lower than those reported in the literature as far as conventional hot smoking is concerned. 27 The above results indicate that the concentration of BaP in the final products was dependent upon the time of processing and pressure conditions. 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