Author’s Accepted Manuscript
Surface hydrophobicity and functional properties of myofibrillar proteins of mantle from frozen stored squid (Illex argentinus) caught either jigging machine or trawling
LorenaA. Mignino, Marcos Crupkin, María E. Paredi
PII: DOI: Reference:
S0023-6438(07)00181-8 doi:10.1016/j.lwt.2007.05.006 YFSTL 1769
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LWT-Food Science and Technology
27 September 2006 27 April 2007 3 May 2007
Cite this article as: Lorena A. Mignino, Marcos Crupkin and María E. Paredi, Surface hydrophobicity and functional properties of myofibrillar proteins of mantle from frozen stored squid (Illex argentinus) caught either jigging machine or trawling, LWT-Food Science and Technology (2007), doi:10.1016/j.lwt.2007.05.006
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1 SURFACE HYDROPHOBICITY AND FUNCTIONAL PROPERTIES OF
2 MYOFIBRILLAR PROTEINS OF MANTLE FROM FROZEN STORED
3 SQUID (Illex argentinus) CAUGHT EITHER JIGGING MACHINE OR
4 TRAWLING 5 Lorena A, Mignino1,2, Marcos Crupkin1,3 and María E. Paredi 1,2,3,*
6
7
1- INTI-Mar del Plata, Marcelo T de Alvear 1168, 7600, Mar del
8
Plata, Argentina.
9 10 11 12 13 14 15 16
2- Comisión de Investigaciones Científicas de la Pcia de Buenos
t Aires (CIC) rip 3- Facultad de Ciencias Agrarias, Universidad Nacional de Mar del
c Plata, Ruta 226, km 73,5, Balcarce, Argentina. nus * Corresponding author. Tel/fax 54-223-4891324/892801 Email: Accepted ma meparedi@mdp.edu.ar
1
1
2 Abstract
3
The surface hydrophobicity and functional properties of actomyosin
4 from mantle of frozen squid caught either by jigging machines (AME1) or 5 by trawl (AME2) were investigated. Two components of 155 and 55 kDa 6 were present in the gels at zero time of storage. Degradation of the 7 myosin heavy chain and increase in the 155 kDa component occur earlier
8 in AME2. Irrespective of the catch method used no significant (p>0.05)
9 10 11 12 13 14 15 16 17 18 19 20
changes in protein solubility were observed. The reduced viscosity of both
t AME1 and AME2 decreased up to months 3 and 5 of frozen storage, rip respectively. At the beginning of storage, the superficial hydrophobicity of c AME2 was 30% higher than that of AME1. SoANS of AME2 significantly us increased during 3 to 5 months of storage period and that of AME1 at the n end of storage. The emulsion activity index (IAE) of AME2 significantly a (p<0.05) increased during the first month and decreased after 3 months of m storage. IAE of AME1 decreased at month 3 and remained unchanged ted thereafter. Emulsion stability (ES) of AME2 showed a behavior that was p similar to its IAE and that of AME1 remained unchanged. Acce Key words: squid, catch method, myofibrillar proteins, functional
21 properties, frozen storage
22
23
2
1 Introduction 2 Illex argentinus is an ommastrephid squid occurring on the continental 3 shelf and slopes of the Southwestern Atlantic Ocean (Roper, Sweeney & 4 Nauen, 1984). It is the most important species of cephalopods in South 5 American waters, according to its potential yield and exportation volume in 6 recent years. About 141,159 tons of squid were caught during 2003 7 (Redes, 2005). Illex argentinus migrates extensively during its life cycle, 8 moving from a presumed spawning area north of the Patagonian shelf to 9 feeding grounds on the shelf, where it grows and reaches sexual maturation
t 10 (Rodhouse and Hatfield, 1990). Mature squids then return to the spawning rip 11 grounds to reproduce and die at the end of one year (Hatakana, 1988). c 12 Squids offer many advantages over other seafood, such as high postus 13 processing yield, very low fat content, bland flavor and very white flesh. In n 14 addition, squid meat has shown to have high functionality which is very a 15 important in food processing. In fish species the functional properties of the m 16 meat, such as water holding capacity, emulsification and gelation capacity, ted 17 are strongly affected by freezing and frozen storage (Sikorski, 1978; p 18 Matsumoto, 1980). These changes are mainly related to modifications in ce 19 myofibrillar proteins (Matsumoto, 1980; Shenouda, 1980). Several authors Ac 20 have reported some aspects related to handling, processing, and frozen
21 storage of squid (Joseph, Varma & Venketaraman, 1977; Botta, Downey, 22 Lauder & Noonan, 1979; Moral, Tejada & Borderias, 1983). A gradual 23 decrease in protein extractability during frozen storage of whole squid Loligo 24 duvauceli (Joseph et al., 1977) and a decrease in extractability, reduced 25 viscosity, and Mg2+-ATPase activity of actomyosin in frozen stored mantles
3
1 of squid (Illex argentinus) (Paredi and Crupkin, 1997) were reported. Similar 2 results were obtained when the same species of squid was frozen stored as 3 whole squid (Paredi, Roldán & Crupkin, 2005). Conversely, it was reported 4 that in other species of squid such as Ommaestrephes sloani pacificus, 5 extractable actomyosin remains without major changes during frozen 6 storage (Iguchi, Tsuchiya & Matsumoto, 1981). 7 The effect of frozen storage on the functional properties of muscle from 8 other squid species was reported (Ruiz-Capillas, Moral, Morales & Montero, 9 2002; Gomez-Guillén, Matinez-Alvarez & Montero. 2003). Ruiz-Capillas et
t 10 al. (2002) observed a decrease in the viscosity and emulsifying capacity of rip 11 protein extracts from mantle and arms of frozen stored squid, either whole or c 12 eviscerated (Illex coindetti). It was also reported that functional properties of us 13 mantle proteins from squid (Loligo vulgaris), remained very stable during n 14 short times of frozen storage (Gómez-Guillén et al., 2003). There are only a a 15 few reports on functional properties of myofibrillar proteins from squid (Illex m 16 argentinus) (Paredi, Davidovich & Crupkin, 1999; Mignino and Paredi, 2006). ted 17 On the other hand, it is widely accepted that the catch method influences the p 18 postmortem biochemical changes in muscle from fish species (Huss, 1995) ce 19 and it had also been reported that when squid was caught by jigging Ac 20 machines a better quality and yield of products, was obtained (Leta, 1989).
21 However, reports on the possible influence of the catch method and frozen 22 storage on the functional properties of myofibrillar proteins from this squid 23 species, are lacking.
4
1 The purpose of the present study was to investigate the behavior of the
2 functional properties of myofibrillar proteins from frozen stored squid
3 harvested by either bottom trawling or jigging machines.
4
5 Materials and methods
6
Squid Illex argentinus (de Castellanos) were harvested by commercial
7 vessels on the Patagonian shelf. Captures were done at 45-52º in the
8 Southwestern Atlantic Ocean. Two experiments were performed. In
9 10 11 12 13 14 15 16 17 18 19 20
experiment 1 (E1) specimens were caught by jigging machines. In
t experiment 2 (E2) specimens were caught by trawl. Ten samples of 10 rip specimens each were packed in polyethylene bags, frozen on board in c blocks at -30ºC and stored at this temperature for 9 months. Frozen samples us were thawed for 12 h at 10°C and six samples of female squid were taken at n zero time (20 days after freezing) and at each period of frozen storage. The a specimens were immediately gutted and after separation of tentacles peeled m off mantles were used for analysis. Only specimens at stage 4-5 (mature) ted were analyzed. The sexual maturation stage of the specimens was p determined according to Brunetti (1990). Acce Actomyosin preparation
21
22
Actomyosin was obtained from mantles according to the method
23 described by Paredi, De Vido de Mattio & Crupkin (1990). The final pellet of
24 actomyosin was solubilized in 0.01m mol/L phosphate buffer (pH 7)
25 containing 0.6 mol/L Na Cl. All the procedures were performed at 0-4ºC.
5
1 Protein determination
2
3
Protein concentrations of actomyosin solutions or protein extracts
4 were determined by the Lowry method, with bovine serum albumin (Sigma
5 Chemical Co., USA) as standard. (Lowry, Rosebrough, Farr & Randall,
6 1951),
7
8 Protein Solubility
9 10 11 12 13 14 15 16 17 18 19 20
t The total myofibrillar extract was obtained by homogenizing 8g of mantle rip (cut into small pieces prior to homogenization) in 160mL of 0.6mol/L KCl c 0.003 mol/L NaHCO3 (pH 7.0) solution for 1 min in a Sorvall Omni-Mixer us 17106 (Dupont Newton, CT, USA) The homogenate was centrifuged for 20 n min at 7500xg in a refrigerated centrifuge Sorvall RC-26 Plus (Sorvall a Product, L.P., Newton, CT, USA) at 2-4 ºC. The supernatant was defined as m the salt soluble protein fraction. Results were expressed as percentage of ted salt-soluble protein respect to total protein determined by the Lowry method p (Lowry et al. 1951). Acce Reduced viscosity
21
22
Reduced viscosity of the actomyosin solution was measured at 20 ±
23 0,1ºC using an Ubbelodhe viscometer (IVA, Buenos Aires, Argentina), by the
24 procedure described by Crupkin, Barassi, Martone & Trucco (1979). The
25 temperature of the viscometer was maintained by a thermostatic bath
6
1 (Thermomix 1480, B. Braun, Germany). Protein concentration covered a
2 range of 0.1-0.4g/100ml.
3
4 Hydrophobicity
5
Protein surface hydrophobicity (So ANS) was determined by the method
6 of Li-Chan, Nakai & Wood (1985). An actomyosin solution (1mg/ml) in 0.010
7 mol/L phosphate buffer (pH6.0) 0.6mol/L KCl was diluted to 0.01-0.05 g of
8 protein per 100 mL using the same buffer. After the temperature was
9 10 11 12 13 14 15 16 17 18 19 20
stabilized at 20ºC, 20µl of 0.008 mol/L1-anilino-8-naphthalene sulfonic acid
t (ANS) in 0.1 mol/L phosphate buffer (pH 7.0) was added to 2mL of diluted rip protein. The relative fluorescence intensity (RFI) values of ANS-conjugates c were measured on a Shimadzu RF-5301PC spectrofluorometer (Kyoto, us Japan) at an excitation wavelength of 370 and an emission wavelength of n 470nm. The initial slope (So) of the RFI versus protein concentration a (expressed as gram of protein per 100mL) plot, calculated by linear m regression analysis, was used as an index of the protein hydrophobicity ted according to the method of Li-Chan et al. (1985). The initial slope is referred p to as So ANS. Acce Emulsifying activity index (EAI) and emulsion stability (ES)
21
22
The emulsions were prepared by the method of Pearce and Kinsella
23 (1978). The actomyosin a 0.1 g/100mL protein solution (w/v, pH 7.0, 3ml)
24 and 1 ml of sunflower oil were homogenized at 5000 rpm for 1 min in a
7
1 Sorvall Omni-Mixer 17106 with microattachment assembly. (Sorvall
2 products, Inc, Newton, CT, USA).
3
EAI and ES were determined by the turbidimetric method of Pearce and
4 Kinsella (1978). The emulsion (50µl) was pipetted from the bottom of the
5 container into 5 ml of 0.1g/100mL sodium dodecyl sulfate (SDS) (w/v)
6 solution, immediately (0min) and 10min after homogenization. Absorbance of
7 the SDS solution was measured at 500nm. Absorbance at 0 time was
8 defined as EAI of protein.
9 10 11 12 13 14 15 16 17 18 19 20
The ES was determined as follows:
t ES= T/T0 rip where T0 and T are turbidities at 0 and 10 min, respectively (Xie & c Hettiarachchy, 1997). The analyses were performed in triplicate. nus SDS-polyacrylamide electrophoresis (SDS-PAGE) ma The SDS-PAGE of actomyosin was performed according to the method ted of Laemmli (1970) using 10g of polyacrylamide per 100g of solution for p separating gel and 4g of polyacrylamide per 100g of solution for the stacking ce gel in a Minislab gel apparatus (Sigma Chemical Co., St Louis, MO, USA). Ac Thirty micrograms of protein were loaded on the gel for each sample, to
21 obtain a linear response with protein concentration. The mobility-molecular
22 weight curve was calibrated with standards of molecular weights (Broad
23 range, BIO-RAD, Bio-Rad Laboratories Inc, Hercules, CA, USA) and
24 contains: rabbit myosin (205 kDa), Escherichia coli β-galactosidase (116
25 .25kDa), rabbit phosphorylase b (97.4 kDa), bovine albumin (66.2 kDa), egg
8
1 albumin (45 kDa), bovine erythrocytes carbonic anhydrase (31 kDa). The
2 voltaje for electrophoresis was set at 90V.
3
Quantitative actomyosin composition was determined by densitometry of
4 the gels at 600nm with a Shimadzu dual-wavelength chromatogram scanner
5 Model CS 910, equipped with a gel scanning accessory (Kyoto, Japan), and
6 the areas of the bands calculated by the triangulation method, as described
7 by Kates (1975). The relative percentages of each band were calculated as
8 follows: (studied band area/Σ of total bands areas) x 100. Myosin/actin and
9 10 11 12 13 14 15 16 17 18 19 20
myosin/paramyosin ratios were calculated by dividing myosin heavy chains
t plus light chain areas by actin and paramyosin areas, respectively (Paredi et rip al., 1990). usc Statistical analysis man Analysis of variance and the Duncan´s new multiple range test were d performed using the Statistica/MAC (Statistica/MAC, 1994) statistical te analysis package. Accep Results and discussion
21 SDS-polyacrylamide electrophoresis (SDS-PAGE)
22
23 SDS-PAGE 10% patterns of actomyosin from mantle of squid harvested by
24 different fishing arts are shown in Fig.1 and Fig. 2. Actomyosin from mantle
25 of squid harvested by either jigging machines (AME1) or trawl shows the
9
1 characteristic polypeptidic bands of myosin heavy chain (MHC), paramyosin
2 (PM), actin (A), tropomyosin (TM), and myosin light chains (MLCs). Similar
3 patterns were reported for actomyosin from this and other species of squid
4 (Iguchi et al., 1981; Paredi & Crupkin, 1997; Mignino & Paredi, 2006). As it
5 can also be seen in Fig. 1 two components of 155 and 55 kDa were also
6 present in the gel of AME1 at zero time and these components remained
7 unchanged up to month 5 of frozen storage. After that, a slight increase in
8 the 155 kDa component and the presence of another one of 143 kDa could
9 10 11 12 13 14 15 16 17 18 19 20
also be observed in the gels. At zero time of storage the SDS-PAGE 10%
t pattern of actomyosin from mantle of squid caught by trawl (E2) also showed rip the presence of both 55 kDa and 155 kDa components (Fig. 2). As it can c also be seen in Fig. 2 a decrease in the MHC band and an increase in 155 us kDa, 104 kDa and 55 kDa bands occur during frozen storage, probably due n to proteolytic activity. a The relative percentages of myosin (M), paramyosin (PM), and actin (A) m and the myosin/actin (M/A) and myosin/paramyosin (M/PM) ratios obtained ted by densitometric analysis of the gels are shown in Table 1. A significant p decrease (p<0.05) in the relative percentage of myosin and in the M/PM ratio ce in AME1, was observed during the last month of frozen storage. A significant Ac decrease (p<0.05) in the M/A ratio in AME1 occurs since month 5 earlier
21 than the decrease in M/PM. Paredi and Crupkin (1997) reported that frozen
22 stored isolated mantles of the same species of squid produce denaturation-
23 aggregation of myofibrillar proteins, especially myosin. In this way, the
24 decrease in the relative percentage of myosin shown in Table 1 could be
25 attributed to denaturation-aggregation of this protein. Conversely, a
10
1 significant decrease (p<0.05) in the relative percentage of myosin and a
2 significant increase (p<0.05) in that of PM, was observed in AME2 since the
3 first month of storage. As a consequence of that, a decrease in both M/A and
4 M/PM ratios, was also observed. Iguchi et al. (1981) reported a decrease in
5 a relative percentage of myosin with an increase in small proteolytic
6 fragments in frozen stored AM from squid (Ommaestrephes sloani pacificus).
7 Cephalopods typically have higher levels of proteolytic activity than most fish
8 species (Kolodziejska & Sikorski; Hurtado, Borderias & Montero, 1999). In
9 10 11 12 13 14 15 16 17 18 19 20
addition, it was reported that myosin was the major target protein for
t proteinases (Nagashima, Ebina, Nakai, Tanaka & Taguchi, 1992; Konno & rip Fukazawa, 1993) and that the proteolytic activity remained unchanged c during the frozen storage (Konno, Young-Je, Yoshioka, Shinho & Seki, us 2003). Konno and Fukazawa (1993) reported that myosin was selectively n cleaved into two large fragments of 150 and 100 kDa which correspond to a heavy and light meromyosin, respectively. In this way, the increase in the m relative percentage of PM shown in Table 1 might be due to commigration of ted this protein with a 104 kDa degradation fragment. Our results suggest that p myosin of AME2 denatured in two steps in mantles of frozen stored squid: ce first myosin is cleaved into 155 and 104 kDa fragments and thereafter the Ac proteolytic fragments aggregate up to the end of storage.
21
22 Protein solubility
23
24
Irrespective of the catch method used no significant changes (p>0.05)
25 in the solubility of protein were observed during frozen storage (Fig. 3). In
11
1 agreement with these results, it was reported that soluble proteins from
2 squid mantle (Loligo vulgaris) remained unchanged after 1 month of frozen
3 storage (Gomez-Guillén et al., 2003) and that protein extractability in frozen
4 stored squids (L. duvaucelli) (Joseph, Perigreen & Nair, 1985) and (O. sloani
5 pacíficus) (Iguchi et al, 1981) only decreases slightly even after long frozen
6 storage. Morales (1997) reported that protein solubility is low sensitive to
7 changes in frozen stored cephalopods muscle.
8
9 10 11 12 13 14 15 16 17 18 19 20
Reduced viscosity and protein surface hydrophobicity
ript Figure 4 shows the changes in reduced viscosity (VER) and surface c hydrophobicity of actomyosin from mantles of frozen stored whole squid. us Viscosity is one of the most sensitive functional properties for measuring n changes in myofibrillar proteins during frozen storage (Barroso, Careche & a Borderias, 1998; Morales, 1997). The reduced viscosity of both AME1 and m AME2 shows a similar behavior up to month 3 of frozen storage. At this time ted of storage a significant (p<0.05) decrease in VER could be observed. p Thereafter, while reduced viscosity of AME1 remained unchanged that of ce AME2 significantly (p<0.05) decreased at month 5 and thereafter remained Ac unchanged. A similar behavior was observed in the reduced viscosity of AM
21 from frozen stored isolated mantles of the same species of squid (Paredi and
22 Crupkin 1997). In addition, a drastic decrease in viscosity of protein extracts
23 during freezing and frozen storage was reported by different authors in
24 different fish species (Mackie 1993; Ruiz Capillas et al., 2002). Several
25 studies on the structure-function relationships in food proteins emphasized
12
1 the importance of protein hydrophobicity on functional properties when
2 different treatments and/or processes were applied (Li-Chan et al, 1985;
3 Nakai, Li-Chan, & Hayakawa, 1986). The aromatic hydrophobicity is widely
4 accepted to monitor changes in the surface hydrophobicity of the proteins 5 (Niwa, Kodha; Kanoh, & Nakayama, 1986; Leblanc & Leblanc, 1992). As it 6 can also be seen in Fig. 4 except for month 3 of frozen storage all the 7 SoANS of AME2 values were higher that those corresponding to AME1.
8 SoANS of AME1 shows a trend to increase between the first and the third
9 10 11 12 13 14 15 16 17 18 19 20
month of frozen storage and remained unchanged thereafter up to month 8.
t A new significant increase (p<0.05) was observed in SoANS of AME1 during rip the last month of storage. SoANS of AME2 remained unchanged up to c month 3 and thereafter showed a trend to increase between months 3 and 5 us of storage and no significant changes were detected thereafter. Niwa et al. n (1986) reported that changes in the hydrophobicity of actomyosin after a freezing are due to myosin rather than to actin. Native myosin has m hydrophobic residues strongly concentrated in the core of the helix ted (McLachlan and Karn, 1982) and the surface of the helix is essentially p devoid of hydrophobic groups (Boredjo, 1983). In this way, the lower surface ce hydrophobicity of AME1 could be due to a greater stability of this protein Ac than that of AME2, suggesting some influence of the catch method on the
21 protein stability.
22
23
24
25
13
1 Emulsifying activity index (EAI) and emulsion stability (ES)
2
3
The changes in IAE of AME1 and AME2 are shown in Fig. 5. At zero
4 time of storage, similar IAE values were observed in both proteins. The IAE 5 of AME2 significantly (p<0.05) increased during the first month of storage 6 and decreased thereafter up to month 7 of storage. No major changes were 7 observed thereafter. The IAE of AME1 significantly (p<0.05) decreased at
8 month 3 and remained unchanged thereafter. At month 1 and 3 of frozen
9 10 11 12 13 14 15 16 17 18 19 20
storage IAE values of AME2 were significantly (p<0.05) higher than those of
t AME1. The higher IAE values of AME2 could be related to proteolytic activity rip detected in AME2 since month 1 of frozen storage. In agreement with our c results a slight increase in the emulsifying capacity of the proteins from us ungutted squid (Illex coindetti) muscle at the beginning of frozen storage n (Ruiz-Capillas et al., 2002) was reported. In that paper, the authors attributed a the increase in the emulsifying capacity to proteolytic activity present in m visceral mass components that penetrated the muscle. Endogenous ted proteolytic activity in mantle of various cephalopods has been described p (Hurtado et al., 1999, Konno and Fukazawa, 1993). In this way, the influence ce of endogenous proteinases of the mantle on the IAE values should not be Ac discarded.
21
The changes in emulsion stability (ES) of AME1 and AME2 are shown
22 in Fig. 6. The ES of AME2 showed a behavior similar to that of IAE.
23 Conversely, the ES values of AME1 remained unchanged up to month 7 and
24 decreased thereafter up to the end of storage. Except for months 5 and 7 of
25 storage ES values of AME1 were lower than those of AME2. Several factors
14
1 have influence on protein stabilized emulsions: rate of diffusion, solubility,
2 viscosity, protein flexibility, net charge, and protein hydrophobicity. In
3 addition, to stabilize an emulsion, a protein must: diffuse to the interface,
4 unfold, expose hydrophobic groups and interact with lipid. In this way, the
5 higher ES values of AME2 respect to AME1 might be due either to a higher
6 unfold and exposition of hydrophobic groups or to a higher content of flexible
7 peptides which can migrate to the interface. In addition, an enhanced
8 emulsion stability of natural actomyosin by apparition of aggregates in the
9
10
11
12 13 14 15 16 17
18
19
20
21
22
extract was reported (Tejada, Mohamed, Huidobro & Garcia, 2003). Further
t investigations will be necessary to clarify the mechanism which led to an rip increase in ES of actomyosin from squid. nusc Conclusion a Actomyosin from squid caught by trawl shows after a short frozen storage m period a higher rate of autolysis, surface hydrophobicity, IAE and ES than d actomyosin from squid harvested by jigging machines. These results indicate te that the catch method influences the rate of autolysis and the functional Accep properties of myofibrillar proteins from frozen stored squid mantle.
15
1 2 3
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8 Moral, A., Tejada, M. & Borderias, A.J. (1983). Frozen storage behaviour of
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squid (Loligo vulgaris) International Journal of Refrigeration, 6, 54- 57.
t Morales, J. (1997) biochemical characterization and behavior in chilled rip storage under control atmospheres or frozen of species: Illex coindetii, c Thoradopsis eblaman and Eledone cirrhosa, PHD Thesis, Univ. us Complutense, Madrid.
n Nagashima, Y., Ebina, H., Nakai, T., Tanaka, M. & Taguchi, T. (1992). a Proteolysis affects thermal gelation of squid mantle muscle. Journal of m Food Science, 57, 916-922. ted Nakai, S., Li-Chan, E. & Hayakawa, S. (1986). Contribution of protein p hydrophobicity to its functionality. Die Nahrung, 30, 327-336. ce 916- 822. Ac Niwa, E., Kodha, S., Kanoh, S. & Nakayama, T. (1986). Exposure of
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hydrophobic amino acid residues from actomyosin on freezing.
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Reconfirmation by fluorometry. Bulletin of Japanese Society Scientific
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Fisheries , 52(6), 1039-1042.
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1 Paredi, M.E., De Vido De Mattio, N. & Crupkin, M. (1990). Biochemical
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4 Paredi, M.E. & Crupkin, M. (1997). Biochemical properties of actomyosin
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from frozen stored mantles of squid (Illex argentinus) at different sexual
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maturation stages Journal of Agricultural and Food Chemistry. 45, 1629-
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8 Paredi, M.E, Davidovich, L.A. & Crupkin, M. (1999). Thermally induce
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gelation of squid (Illex argentinus) actomyosin. Influence of sexual
t maturation stage. Journal of Agricultural and Food Chemistry , 47: rip 3592-3595. c Paredi, M. E., Roldán, H.A. & Crupkin, M. (2005). Effect of frozen storage us on the biochemical properties of actomyosin and lipids composition of n male and female squid (Illex argentinus) mantle. Proceedings of 2nd a Mercosur Congress in Chemical Engineering. 4th Congress on Process m Systems Engineering. Rio de Janeiro, Brasil, ( C6- P36). ted Pearce, K.N., & Kinsella, J.E. (1978) Emulsifying properties of proteins p evaluation of a turbidimetric technique. Journal of Agricultural and Food ce Chemistry., 26: 716-722. Ac REDES, Revista Redes de la Industria Pesquera Argentina. 2005. Nº142,
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maturation in the cephalopods Illex argentinus de Castellanos 1960.
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(Theuthoidea: Ommastrephidae). Philosophical Transactions: Biological
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1 Roper, C.F.E., Sweeney, M.J. & Nauen, C.E. (1984). Cephalopods of the
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world. An annotated and illustrated catalogue of species of interest to
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4 Ruiz- Capillas, C., Moral, A., Morales, J. & Montero, P. (2002). The effect of
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frozen storage on the functional properties of the muscle volador (Illex
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coindetti). Food Chemistry, 78 (2), 148- 156.
7 Shenouda, Y.K. (1980). Theories of protein denaturation during frozen
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storage of fish flesh. Advance in Food Research. 26, 275-311.
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Sikorski, Z.E. (1978). Protein change in muscle food due to freezing and
t frozen storage, International Journal of Refrigeration, 1, 173 -180. rip STATISTICA, MAC. (1994). Statistica for Macintosh; Sttarsoft, Inc. Tulsa,
c Oklahoma us Tejada, M., Mohamed, G. F., Huidobro, A. & Garcia, M.L. (2003) Effect of
n frozen storage of hake, sardine and mixed on natural actomyosin a extracted in salt solutions. Journal of the Science and Food m Agriculture. 83, 1380-1388. ted Xie, Y.R., & Hettiarachchy, N.S. (1997). Xanthan gum effects on solubility p and emulsification properties of soy protein isolate. Journal of Food Acce Science, 62. 1101-1104.
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1 Acknowledgment
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3 The authors would like to thank the Comisión de Investigaciones Científicas 4 de la Provincia de Buenos Aires (CIC), the Universidad Nacional de Mar del 5 Plata (UNMdP) and the Instituto Nacional de Tecnología Industrial (INTI).
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Accepted manuscript
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1 Legends of figures
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3 Figure 1. SDS-PAGE 10% gels of actomyosin from mantle of frozen squid
4 caught by jigging machines (AME1) MHC, myosin heavy chain (200kDa);
5 PM, paramyosin (103kDa); A, actin (45kDa); TM , tropomyosin (36kDa);
6 MLCs, myosin lights chains (18-20kDa). St: Molecular weight markers. 30 µg
7 of protein ( actoymosin) was loaded in each lane of the gel.
8
9 Figure 2. SDS-PAGE 10% gels of actomyosin from frozen stored squid
10 caught by trawl (AME2) MHC, myosin heavy chain (200kDa); PM,
11 paramyosin (103kDa); A, actin (45kDa); TM , tropomyosin (36kDa); MLCs,
12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27
myosin lights chains (18-20kDa). St: Molecular weight markers. 30 µg of
t protein ( actoymosin) was loaded in each lane of the gel. crip Figure 3. Changes in solubility of protein of squid mantle during storage at – s 30ºC. Experiment 1 ( ); Experiment 2 ( ). Results are expressed as the u means of 6 determinations ± SD. an Figure 4. Surface hydrophobicity (SoANS): () and Reduced viscosity m (VER): ( ∆ ) of actomyosin from squid mantle during storage at –30ºC. Open d symbols (AME1), closed symbols indicated (AME2). Results are expressed te as the means of 6 determinations ± SD. ep Figure 5. IAE of actomyosin from squid mantle during storage at –30ºC. c Results are expressed as the means of 4-6 determinations ± SD. AME1 ( ) Ac ; AME2 ( ). a,b,c,d,e. It represents a significant difference (p<0.05) in data
28 from different months and same experiment.
29 * Indicate significant differences (p < 0.05) between experiments within same
30 month.
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32
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1 Figure 6 . ES of actomyosin from squid mantle during storage at –30ºC.
2 Results are expressed as the means of 4-6 determinations ± SD. AME1( ); 3 Experiment 2 ( ). 4 a,b,c,d,e . It represents a significant difference (p<0.05) in data from different
5 months and same experiment. 6 * Indicate significant differences (p < 0.05) between experiments within same
7 month
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Table 1. Relative percentage of myosin (M), actin (A) and paramyosin (PM) and
11 M/A, M/PM ratio of actomyosin from squid mantle during frozen storage.
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Relative percentage(%)ª
Ratioª
Time
A
PM
M/A
M/PM
M
0 (E1) 0 (E2)
51.17±8.3b,x 43.35±6.2b,x
27.78±7.9b,x 23.65±2.5b,x
10.73±4.3b,x 15.67±2.3b,x
1.88±0.8b,x 1.86±0.4b,x
5.06±2.8b,x 3.05±0.7b,x
ipt 1 (E1) r 1 (E2)
49.13±1.3b,y 23.36±2.2c,x
24.83±4.1b,x 27.89±4.3b,x
8.97±3.2b,y 22.22±1.8c,x
2.06±0.73b,y 0.85±0.3c,x
4.11±0.25b,y 1.03±0.1c,x
sc 3 (E1) u 3 (E2)
51.30±2.8b,y 16.98±2.8c,x
30.66±6.3b,x 36.58±3.8c,x
10.33±2.2b,y 24.31±2.8c,x
1.73±0.4b,y 0.48±0.1c,x
5.09±1.0b,x 0.72±0.2c,x
n 5 (E1) a 5 (E2)
48.14±4.0b,y 16.93±2.2c,x
33.60±4.5c,x 37.35±2.0c,x
9.30±3.0b,y 29.63±1.8c,x
1.44±0.2c,x 0.45±0.2c,x
3.98±0.1b,y 0.57±0.2c,x
m 7 (E1) d 7 (E2)
42.82±6.8b,y 20.46± 8.2c,x
32.50±8.2c,x 30.90±1.4c,x
8.20±4.4b,y 24.30±6.0c,x
1.40±0.4c,x 0.59±0.2c,x
5.15±1.5b,y 0.86±0.6c,x
te 9 (E1) p 9 (E2)
32.65±6.7c,y 5.20±1.6d,x
43.77±9.1cx 34.07±3.8c,x
10.88±3.4b,y 30.32±3.5c,x
0.87±0.05c,x 0.15±0.5c,x
2.44±0.3b,x 0.20±0.06c,x
ce ª Each v1a4lue represents the mean ± SD (n=4-6). c b,c,d Mea1n5s within each column with different superscrips were significantly different (p<0.05) within sample during frozen storage.
x,y Mea1n6s within each column with different superscrips were significantly different (p<0.05) within sample different experiment,
A same tim1e7of storage at –30ºC.
E1: Expe1r8iment with squid cacth by jiggins machine, E2: Experiment with squid cacth by botton trawl.
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