A method to prepare fish protein hydrolysate from miscellaneous fish obtained as by catch from shrimp trawlers is outlined. Effect of temperature and concentration of enzyme papain on the yield of hydrolysates has been determined. It is seen that within 30 min at 55°C and pH 6.5 fish proteins can be effectively solubilized, provided the nitrogen content of the enzyme (activity 10 units/mg enzyme) and substrate are maintained in the ratio 1:30. This hydrolysate possesses the best amino acid pattern compared to those obtained after hydrolysis for 60 to 180 min

Gopakumar, K.

James, M.A.

Nair, A.L.

Shenoy, A.V.

Thankamma, R.

Aquatic Commons

Fish. Techno!. (1979) 16, 71-75 71 
Protein Hydrolysate ft'om Miscellaneous Fish 
R. THANKAMMA, K. GOPAKUMAR, A. LEKSHMY NAIR, 
A. V ASANTH SHENOY AND M. ARUL JAMES 
Central Institute of Fisheries Technology, Cochin-682 029 
A method to prepare fish protein hydrolysate from miscellaneous fish obtained as by 
catch from shrimp trawlers is outlined. Effect of temperature and concentration 
of enzyme papain on the yield of hydrolysates has been determined. It is seen that 
within 30 min at 55°C and pH 6.5 fish proteins can be effectively solubilised, provided 
the nitrogen content of the enzyme (activity 10 units/mg enzyme) and substrate 
are maintained in the ratio 1 :30. This hydrolysate possesses the best amino acid 
pattern compared to those obtained after hydrolysis for 60 to 180 min. 
Miscellaneous fish constitute a major 
portion of the fish catch. At present they 
are not utilised fully and some attempts 
have been made to utilise them (Anon, 1973, 
'76, '77, '78). Protein hydrolysate from mis-
cellaneous fish (trash fish) has several 
commercial applications. Dietary protein 
is absorbed in the form of oligo peptides 
and free amino acids. Protein hydro-
lysates which contain oligo peptides and 
free amino acids have better physiological 
advantages in correcting malnutrition com-
pared to a mixture of free amino acids 
(Mathews, 1971; Silk, 1974). 
Fish protein has been used as raw 
material for the production of hydrolysates 
by many workers. The factors affecting 
the degree of hydrolysis and the fractional 
analysis of certain papaic hydrolysates of 
fish flesh have been reported by Sen et a!. 
(1962) and Sripathy et a!. (1962). Sripathy 
et a!. (1963) showed that fish hydrolysates 
are of good nutritional value. Technolo-
gical aspects relating to the production of 
hydrolysate from a fresh water fish, Barbus 
dubius using papain have been worked 
out by Sripathy et a!. (1964). Hale (1969) 
measured the relative activities of more 
than twenty commercially available pro-
teolytic enzymes for the digestion of a 
washed and freeze-dried fish protein subs-
trate and showed that pepsin, papain and 
pancreatin all combined good activity 
with moderate cost. Hale (1972) has also 
reported effects of various processing con-
ditions and commerically available proteo-
lytic enzymes on yield and composition 
Vol. 16 
of water soluble fish protein hydrolysates. 
According to McBride et al. (1961) pepsin 
produced better solubilization of herring 
tissue than other proteolytic enzymes. 
Yanez et a!. (1976) employed hake as raw 
material for the production of protein 
hydrolysates and used it for the supple-
mentation of cereal protein. However, one 
of the disadvantages of protein hydrolysate 
is its bitterness,. caused by the presence of 
bitter peptides. Recently, Lalasidis et a!. 
(1978) and Lalasidis & Sjoberg (1978) 
used a powerful bacterial endopeptidase 
alcalase, followed by pancreatin to produce 
bitter-free protein hydrolysate from deboned 
cod filleting offal, fish protein concentrate 
and fresh herring. Both pepsin and 
papain were initially tried to produce hydro-
lysates by the authors. Subsequently 
pepsin was discontinued from the studies 
as it involved hydrolysis at a lower pH, 
which necessitated . the neutralisation of 
the final product thereby increasing 
concentration of salt in the finished pro-
duct. The physical properties of peanut 
flour hydrolysed by bromelin, pepsin and 
trypsin have been reported by Beuchat 
et a!. (1975), but pepsin and trypsin are too 
expensive for commerical applications. 
Papain is reported to be more desirable in 
proteolysis than bromelin (Sekul & Ory, 
1977). Sekul et al. (1978) also showed 
that proteolysis of peanut proteins with 
papain caused certain functional changes 
in the properties of hydrolysed proteins 
with certain advantages in food applica-
tions. The present study reports the effect 
of temperature, time and concentration of 
72 R.THANKAMMA,K.GOPAKUMAR,A.L. NAIR,A.V. SHENOY ANDM.A. JAMES 
enzyme on the yield of protein hydroly-
sate from miscellaneous fish. 
Materials an1l Methods 
Miscellaneous fish were obtained from 
shrimp trawlers operated off Cochin. 
Commercial grade papain (activity 10 units/ 
mg enzyme) was employed. Quantity of 
the enzyme required to hydrolyse fish 
proteins was calculated in terms of the 
ratios of the nitrogen contents of both enzyme 
and substrate, following the method of 
Nanda et al. (1977). The quantity of 
minced fish meat used varied from 1-15 kg. 
The comminuted whole fish and picked 
fish meat were separately mixed with 1 ;1 
wfw of water, sterilized at 0.7 kg/sq em 
pressure for 30 min and cooled. The pH 
was adjusted to 6. 5 with hydrochloric acid 
and transferred to a reaction vessel main-
tained at different temperatures with the 
help of a thermostatic water bath. At 
these temperatures the required quantity 
of enzyme dissolved in minimum quantity 
of distilled water was added and the reaction 
mixture stirred continuously. After specific 
intervals of time varying from 15 to 180 
min the reaction was stopped by boiling 
the contents, filtered through Whatman 
No. 1 filter paper and the hydrolysate 
separated, pH adjusted to 7, concentrated· 
on a water bath and dried under vacuum. 
The amino acid composition of the protein 
hydrolysates prepared from threadfin bream 
was estimated by microbiological assay 
methods of Kavanagh (1963). 
Results and Discussion 
Yield of protein hydrolysates at diff-
erent concentrations of enzyme is given in 
Table 1. The effect of time and temperature 
on the yield of hydrolysate is presented in 
Tables 2 and 3. Table 4 gives the amino 
acid composition of protein hydrolysates 
prepared from threadfin bream hydrolysed 
for different periods using same concen-
tration of enzyme (enzyme nitrogen to 
substrate nitrogen ratio 1 :30). Yields of 
protein hydrolysates from different species 
of trash fish are given in Table 5. 
Table 1 shows that among the differ-
ent concentrations of papain used to hydro-
lyse fish proteins the optimum yield is 
obtained when enzyme and substrate 
Table 1. Yield* of fish protein hydrolysate with respect to change in enzyme (papain) 
concentration (pH 6.5, temperature 55oC and time 180 min) 
Fish Ratio of enzyme nitrogen to substrate nitrogen 
1 :20 1 :30 1 :40 1 :60 
Throadfin bream (whole) Sample 1 
Sample 2 
Threadfin bream meat (picked) 
Jew fish (whole) 
* g/100 g whole fish/picked meat 
12.4 
12.1 
7.9 
8.9 
11.3 
11.0 
7.8 
8.0 
8.9 
10.2 
7.4 
6.1 
Table 2. Yield* of fish protein hydrolysate with respect to time of hydrolysis 
(enzyme nitrogen to substrate nitrogen ratio 1 :30, pH 6.5, temperature 55°C) 
Time in min 
Fish 15 30 60 120 180 
Threadfin bream (whole) Sample 1 8.8 10.2 10.2 10.5 10.6 
Sample2 10.9 11.4 11.5 12.2 12.2 
Threadfin bream meat (picked) 9.0 9.6 10.0 10.2 10.0 
Sole (whole) 11.2 11.8 11.8 12.3 12.3 
Glassy perchlet (whole) 10.0 10.7 10.8 11.2 11.3 
* g/100g whole fish/picked meat 
7.3 
10.2 
7.1 
5.3 
1979 
PROTEIN HYDROLYSATE FROM MISCELLANEOUS FISH 73 
Table 3. Effect of temperature on hydrolysis of minced whole fish 
(enzyme nitrogen to substrate nitrogen ratio 1 :30, pH 6.5, time of hydrolysis 30 min) 
Yield* of hydrolysate at 
Sample 40°C 50°C 55°C 60°C 70oc 
1 9.0 10.8 11.3 10.7 10.0 
2 10.7 11.5 12.9 11.3 10.7 
3 10.5 12.2 12.8 12.4 12.3 
4 8.7 8.8 11.1 9.3 8.9 
':' gj I OOg whole fish 
Table 4. Amino acid composition of protein hydrolysates at different times of hydrolysis 
(enzyme nitrogen to substrate nitrogen mtio 1 :30, pH 6.5, temperature 55°C) 
Amino acids* 
30min 
Lysine 7.32 
Methionine 2.67 
Leucine 3.03 
Isoleucine 2.36 
Threonine 5.11 
Tryptophan 0.99 
Phenylalanine 4.43 
Valine 2.26 
Arginine 4.25 
Glutamic acid 10.51 
Aspartic acid 4.62 
Histidine 2.51 
Glycine 2.50 
Serine 4.75 
Proline 3.61 
Cystine 0.47 
Tyrosine 4.81 
* g/lOOg protein 
nitrogen were in the ratio I :30. Using 
this ratio, the effect of reaction time on 
proteolysis was worked. out (Table 2). 
Though maximum yield of hydrolysate 
was obtained when hydrolysis extended 
up to ISO min the differences in yield was 
not appreciable compared to that at 30 min. 
Further reduction in time of hydrolysis 
to 15 min entailed in certain cases appre-
ciable lowering in the yield of hydrolysate, 
suggesting that a viable yield could be 
obtained after 30 min hydrolysis. 
The study of the effect of temperature 
on proteolysis of fish proteins with enzyme 
Vol. 16 
Hydrolysate 
60min 120 min 180 min 
8.34 7.10 7.92 
2.54 2.71 3.20 
2.14 3.16 3.21 
3.14 3.43 3.11 
4.96 4.93 4.87 
0.89 0.79 0.84 
4.39 4.41 4.06 
1.96 2.43 2.37 
5.31 5.52 5.67 
13.56 13.60 13.41 
4.91 5.26 5.71 
3.91 5.73 4.97 
2.91 2.46 2.62 
5.25 4.95 4.31 
4.66 4.21 3.95 
0.63 0.62 0.69 
5.24 5.40 5.16 
papain shows that (Table 3) an operational 
temperature of 55°C will be the most suitable 
for the maximum yield of hydrolysate. 
The study of the overall amino acid 
pattern of the different hydrolysates produ-
ced at 30, 60, 120 and 180 min showed very 
little variation in the individual amino 
acid make up. However, it should be 
noted that tryptophan, phenylalanine and 
threonine are lost significantly when hydro-
lysis was extended to ISO min. Glycine, 
proline, leucine and valine showed very 
little change between 30 and 180 min. 
The rest of the amino acids showed a 
74 R. THANKAMMA, K. GOPAKUMAR,A. L. NAIR, A. V. SHENOY AND M.A. JAMES 
Table 5. Yield of protein hydrolysate from miscellaneous fish 
(enzyme nitrogen to substrate nitrogen ratio 1 :30, pH 6.5, temperature 55°C, time of 
hydrolysis 30 min). 
Common name Species name Yield':' of hydrolysate 
Lizard fish 
Large spined flat head 
Ribbon fish 
Saurida tumbil 
Platycephalus macracanthus 
Trichiurus sp. 
13.3 
11.0 
9.9 
11.9 
9.9 
12.0 
10.9 
9.7 
Barracuda Sphyraena sp. 
Jew fish Jolmius sp. 
Threadfin bream 
Cat fish 
Nemipterus japonicus 
Tachysurus sp. 
Anchovies Thrissocles sp. 
Sole Cynoglossus sp. 8.6 
'~ g/lOOg whole fish 
marginal increase. Considering the loss 
of tryptophan, phenylalanine and 
tlu·eonine (essential amino acids) it is not 
advisable to carry out hydrolysis up to 
180 min from the nutritional point, as they 
will become limiting amino acids when 
protein scores are evaluated to incorporate 
hydrolysates in food preparations. It is 
also reported that extended enzymatic 
digestion of fish proteins causes a reduction 
of their nutritional .value and a partial 
destruction of essential amino acids (Gillies, 
1975). Thus the best amino acid pattern 
is . obtained in the product after 30 min 
hydrolysis. 
Picked fish meat, obtained with the 
help of a mechanical flesh separator, which 
removes bones, scales and skin from meat, 
invariably gave lower yield compared to 
whole fish meat. The higher yield of 
hydrolysates from whole fish can be inter-
preted as due to the presence of calcium 
ions in the meat, obtained from bones, 
which is found to activate papain. 
The authors are indebted to Shri G. K. Kuriyan, 
Director, Central Institute of Fisheries Technology 
and Shri M. Rajendranathan Nair, Project Co-ordi-
nator for permission to publish the paper. 
References 
Anon (1973) Proc. ii Workshop, Indian 
Council of Agricultural Research Co-
ordinated Research Project on Transport-
ation of Fresh Fish and Utilization of 
Trash Fish. Mangalore, April 5-8 
(unpublished) 
Anon (1976) Proc. IV Workshop, Indian 
Council of Agricultural Research Co-
ordinated Research Project on Trans-
portation of Fresh Fish and Utilization 
of Trash Fish. Cochin, October 27-28 
(unpublished) 
Anon (1977) Proc. V Workshop,. Indian 
Council of Agricultural Research Co-
ordinated Research Project on Trans-
portation of Fresh Fish and Utlization 
of Trash Fish. Madras, November 
15-16 (unpublished) 
Anon (1978) Proc. VI Workshop, Indian 
Council of Agricultural Research Co-
ordinated Research Project on Trans-
portation of Fresh Fish and Utilization 
of Trash Fish. Mangalore, November 
3-4 (unpublished) 
Beuchat, L.R., Cherry, J.P. & Quinn, M.R. 
(1975) J. agric. Fd chem. 23, 616 
Gillies, M. T. (1975) Fd Techno!. Rev. 
22, 313 
Hale, M.B. (1969) Fd Techno!. 23, 107 
Hale, M.B. (1972) NOAA Technical 
Report NMFS SSRD 657, Making 
Fish Protein Concelltrates by Enzymatic 
Hydrolysis 
1979 
PROTEIN HYDROLYSATE FROM MISCELLANEOUS FISH 75 
Kavanagh, F. (1963) In Analytical 
Microbiology, 1st edn., p. 167, Acade-
mic press, New York 
Lalasidis, G., Bostrom, S. & Sjoberg, L.B. 
(1978) J. agric. Fd chem. 26, 751 
Lalasidis, G. & Sjoberg, L.B. (1978) J. 
agric. Fd chem. 26, 742 
Mathews, D.M. (1971) J. Clin. Pathol. 
24, Suppl. (Roy. Col!. Path) 5, 29 
McBride, J.R., Idler, D.R. & Macleod, 
R.A. (1961) J. Fish. Res. Bd Can. 
18, 93 
Nanda, C.L., Ternmouth, J.H. & Kondas, 
A.C. (1977) J. Sci. Fd Agric. 28, 20 
Sekul, A.A. & Ory, R.L. (1977) J. Am. 
Oil chem. Soc. 54, 32 
Vol. 16 
Sekul, A.A., Vinnet, C.H. & Ory, R.L. 
(1978) J. agric. Fd chem. 26, 855 
Sen, D.P., Sripathy, N.V., Lahiry, N.L., 
Sreenivasan, A. & Subramanyam, V. 
(1962) Fd Techno!. 16, 138 
Silk, D.B.D. (1974) Gut. 15, 494 
Sripathy, N.V., Sen, D.P., Lahiry, N.L., 
Sreenivasan, A. & Subramanyam, V. 
(1962) Fd Techno!. 16, 141 
Sripathy, N.V., Kadkol, S.B., Sen, D.P., 
Swaminathan, M. & Lahiry, N.L. 
(1963) J. Fd Sci. 28, 365 
Sripathy, N.V., Sen, D.P. & Lahiry, N.L. 
(1964) Res. & Ind. 9, 258 
Yanez, E., Ballester, D. & Monckeberg, F. 
(1976) J. Fd Sci. 41, 1289 


Protein hydrolysate from miscellaneous fish

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Protein hydrolysate from miscellaneous fish

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