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Original

Prev Nutr Food Sci 2017; 22(2): 90-99

Published online June 30, 2017 https://doi.org/10.3746/pnf.2017.22.2.90

Copyright © The Korean Society of Food Science and Nutrition.

Differential Effects of Cod Proteins and Tuna Proteins on Serumand Liver Lipid Profiles in Rats Fed Non-Cholesterol- and Cholesterol-Containing Diets

Ryota Hosomi1, Hayato Maeda2, Yuki Ikeda1, Yuko Toda1, Munehiro Yoshida1, and Kenji Fukunaga1

1Laboratory of Food and Nutritional Sciences, Faculty of Chemistry, Materials, and Bioengineering, Kansai University, Osaka 564-8680, Japan 2Laboratory of Food Chemistry, Faculty of Agriculture and Life Science, Hirosaki University, Aomori 036-8561, Japan

Received: February 6, 2017; Accepted: March 17, 2017

This is an Open Access article distributed under the terms of the Creative Commons Attribution Non-Commercial License (http://creativecommons.org/licenses/by-nc/4.0) which permits unrestricted non-commercial use, distribution, and reproduction in any medium, provided the original work is properly cited.

Abstract

ABSTRACT: Fish muscles are classified into white and red muscles, and the chemical composition of the two fish muscles have many differences. Few reports have assessed the health-promoting functions of white fish muscle proteins (WFP) and red fish muscle proteins (RFP). We therefore evaluated the mechanisms underlying the alteration of lipid profiles and cholesterol metabolism following the intake of WFP prepared from cod and RFP prepared from light muscles of tuna.Male Wistar rats were divided into six dietary groups: casein (23%), WFP (23%), and RFP (23%), with or without 0.5% cholesterol and 0.1% sodium cholate. Compared to the WFP-containing diet, the RFP-containing diet supplemented with cholesterol and sodium cholate significantly increased serum and liver cholesterol contents. However, in the RFP groups, an alteration in cholesterol metabolism including an increased tendency to excrete fecal sterols and hepatic cholesterol 7α-hydroxylase was related to the reduction of hepatic cholesterol contents. This phenomenon might be related to the tendency of an increased food intake in RFP-containing diets. These results highlight the differential effects of WFP and RFP on serum and liver lipid profiles of Wistar rats fed non-cholesterol- or cholesterol-containing diets under no fasting condition.

Keywords: cod protein, tuna protein, lipid metabolism, cholesterol, rat

Fig 1.

Figure 1.

Sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) patterns of casein, WFP, and RFP. SDS-PAGE was performed using 12.5% polyacrylamide gels. M, molecular weight marker; CAS, casein; WFP, white muscle fish protein; RFP, red muscle fish protein.

Preventive Nutrition and Food Science 2017; 22: 90-99https://doi.org/10.3746/pnf.2017.22.2.90

Fig 2.

Figure 2.

Time courses of degree of hydrolysis and insoluble fraction production rate in simulated gastrointestinal digestions. Data represent means±SEM (n=4). (A) Significant effects of time, protein, and time-protein interactions were identified by two-factor repeated measure analysis of variance. (B) Values in the same row not sharing a common letters (a,b) are significantly different at P<0.05 using the Tukey’s multiple comparisons test. RFP, red muscle fish protein; WFP, white muscle fish protein.

Preventive Nutrition and Food Science 2017; 22: 90-99https://doi.org/10.3746/pnf.2017.22.2.90

Table 1 . Composition of the experimental diets (unit: g/kg).

Experiment 1Experiment 2


CASWFPRFPCAS+CWFP+CRFP+C
Casein230230
WFP230230
RFP230230
Dextrinized corn starch92.192.192.192.192.192.1
Corn starch277.386277.386277.386271.386271.386271.386
Sucrose100100100100100100
Cellulose505050505050
AIN-93G mineral mixture353535353535
AIN-93 vitamin mixture101010101010
L-Cystine333333
Choline bitartrate2.52.52.52.52.52.5
Soybean oil707070707070
Lard130130130130130130
Cholesterol555
Sodium cholate111
tert-Butylhydroquinone0.0140.0140.0140.0140.0140.014

CAS, casein; WFP, white muscle fish protein; RFP, red muscle fish protein; CAS+C, casein with cholesterol; WFP+C, WFP with cholesterol; RFP+C, RFP with cholesterol; AIN, American Institute of Nutrition..


Table 2 . Chemical composition of casein, WFP, and RFP.

ComponentCaseinWFPRFP
Crude protein (g/100 g)85.985.485.7
 Amino acid compositions (wt %)
  Alanine2.36.36.6
  Arginine3.68.07.2
  Aspartic acid1)4.910.19.8
  Glutamic acid2)18.516.614.5
  Glycine1.54.54.2
  Histidine2.52.68.2
  Isoleucine4.44.14.3
  Leucine8.37.97.5
  Lysine12.09.47.1
  Methionine2.33.23.3
  Phenylalanine5.54.13.8
  Proline13.64.63.5
  Serine4.44.53.9
  Threonine3.94.74.8
  Tyrosine6.53.73.5
  Valine5.74.54.4
 Mb (g/100 g)ND0.10.4
Crude fat (g/100 g)0.71.00.7
 EPA+DHA (g/100 g)ND0.20.1
 Cholesterol (mg/100 g)20.010.013.0
Moisture (g/100 g)5.65.95.3
Ash (g/100 g)1.86.35.7

WFP, white muscle fish protein; RFP, red muscle fish protein..

1)Aspartic acid+asparagine.

2)Glutamic acid+glutamine.

EPA, eicosapentaenoic acid; DHA, docosahexaenoic acid; Mb, myoglobin; ND, not detected..


Table 3 . Growth parameters and relative organ weights in rats fed experimental diets for 4 weeks.

Experiment 1Experiment 2


CASWFPRFPCAS+CWFP+CRFP+C
Growth parameters
 Initial BW (g)72.9±1.171.9±0.971.8±1.184.6±2.583.5±1.984.5±2.3
 Final BW (g)303.6±10.8305.1±4.5320.2±3.9317.1±8.3322.9±5.4344.2±10.4
 BW gain (g/d)8.2±0.48.3±0.28.9±0.18.6±0.48.9±0.29.6±0.3
 Food intake (g/d)17.3±0.817.3±0.819.6±0.917.9±0.717.7±0.720.0±0.9
 Food efficiency (g/g)0.48±0.020.48±0.010.45±0.010.48±0.020.50±0.010.48±0.02
 Water intake (mL/d)35.7±2.636.0±2.737.4±2.833.5±2.132.5±2.034.0±2.3
Organ weight (g/100 g BW)
 Liver weight4.43±0.324.20±0.084.23±0.066.07±0.11b5.32±0.13a6.18±0.21b
 Perirenal WAT weight1.28±0.141.32±0.101.46±0.191.36±0.091.35±0.111.80±0.26
 Mesentery WAT weight1.26±0.201.36±0.081.34±0.181.51±0.171.57±0.061.58±0.08
 Epididymal WAT weight1.33±0.151.42±0.071.41±0.051.57±0.151.37±0.171.54±0.24

Data represent means±SEM (n=6)..

Values in the same row not sharing a common letters (a,b) are significantly different at P<0.05 using the Tukey’s multiple comparisons test..

CAS, casein; WFP, white muscle fish protein; RFP, red muscle fish protein; CAS+C, casein with cholesterol; WFP+C, WFP with cholesterol; RFP+C, RFP with cholesterol; BW, body weight; WAT, white adipose tissue..


Table 4 . Biochemical parameters in serum, liver, and feces in rats fed experimental diets for 4 weeks.

Experiment 1Experiment 2


CASWFPRFPCAS+CWFP+CRFP+C
Serum biochemical parameters
 AST (IU/L)73.7±3.869.2±1.072.3±1.465.7±3.665.2±2.569.0±3.0
 ALT (IU/L)42.8±3.040.8±1.738.0±1.334.8±3.539.5±2.634.7±2.8
 TAG (mg/dL)130.8±23.7116.4±26.079.4±12.1162.5±28.4164.2±29.9181.5±29.9
 Cholesterol (mg/dL)82.5±6.084.5±5.694.3±2.894.5±1.3a85.0±3.5a114.3±6.1b
 HDL-cholesterol (mg/dL)55.5±5.358.8±4.267.5±2.552.8±2.2ab48.0±2.2a63.2±3.6b
 Non-HDL-cholesterol (mg/dL)27.0±1.025.7±1.426.8±0.444.0±2.5ab37.0±1.8a51.2±3.8b
 PL (mg/dL)149.5±9.8151.7±5.2168.2±3.5156.3±7.2a144.8±2.6a187.0±7.1b
 NEFA (μEq/L)1,036.3±133.1926.5±62.0978.6±197.4653.8±33.3ab551.0±12.3a748.2±56.4b
Liver lipid contents (mg/g)
 TAG78.7±9.5AB52.8±3.8A111.8±18.9B195.7±11.3168.6±13.1192.2±19.3
 Cholesterol4.11±0.48B2.51±0.36A3.26±0.31B6.79±0.82a4.76±0.45a12.93±1.11b
 PL21.2±1.220.0±0.619.2±1.321.7±0.822.0±0.420.0±0.8
Fecal biochemical parameters
 Dry weight (g/d)7.77±0.248.16±0.148.61±0.316.10±0.338.88±1.48.09±0.52
 FFA (mg/d)124.0±8.4138.1±4.7141.2±10.5316.7±32.8435.4±61.1414.9±65.7
 Neutral sterols (mg/d)14.2±1.317.7±2.718.8±1.637.5±2.872.4±16.059.8±8.0
 Acidic sterols (mg/d)13.6±2.3A16.7±2.0AB21.9±1.6B22.0±2.2a35.2±3.8ab39.9±6.0b
 Nitrogen content (mg/d)9.36±0.56A10.21±0.27A13.22±0.70B6.05±0.90a11.32±2.42ab13.57±1.13b

Data represent means±SEM (n=6)..

Values in the same row not sharing a common letters (a,b or A,B) are significantly different at P<0.05 using the Tukey’s multiple comparisons test..

CAS, casein; WFP, white muscle fish protein; RFP, red muscle fish protein; CAS+C, casein with cholesterol; WFP+C, WFP with cholesterol; RFP+C, RFP with cholesterol; AST, aspartate aminotransferase; ALT, alanine aminotransferase; TAG, triacylglycerol; HDL, high-density lipoprotein; PL, phospholipid; NEFA, non-esterified fatty acid; FFA, free fatty acid..


Table 5 . Enzymes activities and relative mRNA expression levels in livers from rats fed experimental diets for 4 weeks.

Experiment 1Experiment 2


CASWFPRFPCAS+CWFP+CRFP+C
Enzyme activity (nmol/min/mg protein)
 CPT-21.46±0.231.38±0.441.32±0.221.37±0.081.15±0.460.70±0.16
 ACOX2.17±0.473.06±0.322.23±0.601.90±0.12b1.72±0.24ab1.08±0.15a
 FAS1.39±0.17AB1.20±0.15A1.97±0.21B1.35±0.131.26±0.051.25±0.07
 ACC56.8±5.2A55.8±8.2A91.7±10.1B66.5±4.884.6±10.997.6±5.4
 G6PDH10.8±0.9A12.5±1.7AB24.4±5.4B4.5±0.2a3.8±0.2a7.5±1.1b
 ME9.7±1.1A16.2±.4AB21.8±2.2B4.2±0.34.4±0.34.9±0.2
mRNA expression level (arbitrary units)
 ABCA1100.0±23.096.6±23.882.5±20.9100.0±16.585.1±24.1100.2±48.4
 ABCG5100.0±21.2102.8±16.6101.1±17.8100.0±11.989.5±13.991.8±16.4
 ABCG8100.0±28.7153.4±20.5182.3±66.2100.0±16.155.9±24.141.5±14.0
 ACAT-1100.0±2.892.2±12.770.6±13.6100.0±18.792.7±27.5106.3±48.3
 CYP7A1100.0±19.0161.6±45.891.7±31.4100.0±17.9201.4±36.0213.9±35.2
 HMGCR100.0±7.3115.6±29.8106.2±13.9100.0±12.558.4±17.686.6±15.7
 LDLR100.0±19.659.6±15.061.2±12.1100.0±22.662.3±8.151.6±7.2
 SRB1100.0±24.666.1±9.344.2±12.3100.0±36.854.0±15.171.4±19.3
 SHP-1100.0±21.4158.7±50.0147.3±72.6100.0±19.050.3±11.050.1±9.8
 SREBF-2100.0±22.264.5±17.759.1±10.4100.0±8.261.1±19.375.2±15.3

Data represent means±SEM (n=6)..

Values in the same row not sharing a common letters (a,b or A,B) are significantly different at P<0.05 using the Tukey’s multiple comparisons test..

The mRNA expression levels were determined by real-time polymerase chain reaction analysis using the glyceraldehyde 3-phosphate dehydrogenase mRNA expression level for normalization. mRNA expression levels of genes are shown relative to those determined from livers of rats fed the CAS (Experimental 1) and CAS+C (Experimental 2) diets (set at 100)..

CAS, casein; WFP, white muscle fish protein; RFP, red muscle fish protein; CAS+C, casein with cholesterol; WFP+C, WFP with cholesterol; RFP+C, RFP with cholesterol; CPT-2, carnitine palmitoyltransferase-2 ACOX, acetyl-CoA oxidase; FAS, fatty acid synthase; ACC, acetyl-CoA carboxylase; G6PDH, glucose-6-phosphate dehydrogenase; ME, malic enzyme; ABCA1, ATP-binding cassette subfamily A1 ABCG, ATP-binding cassette subfamily G ACAT-1, acetyl-CoA acetyltransferase-1 CYP7A1, cholesterol 7α-hydroxylase; HMGCR, 3-hydroxy-3-methyl-glutaryl-CoA reductase; LDLR, low-density lipoprotein receptor; SRB1, scavenger receptor class B member 1; SHP-1, small heterodimer partner-1; SREBF-2, sterol regulatory element binding factor-2..