Pertanika J. Trop. Agric. Sci. 19(1): 69-75 (1996)ISSN: 0126-6128

Penerbit Universiti Pertanian Malaysia

Differences in Functional Properties of Mungbean Protein Concentrate andthe Effect of Incorporation into Fish Sausages

SUHAILA MOHAMED*, JAMILAH BAKAR and NORHASHIMAH ABD HAMIDFaculty of Food Science and Biotechnology

Universiti Pertanian Malaysia43400 UPM Serdang, Selangor Darul Ehsan, Malaysia

Keywords: emulsion, fat absorption, foaming, fish sausages, mungbean concentrate

ABSTRAKSifat fisiko'kimia dan sifat berfungsi pekatan protein kacang hijau yang dihasilkan melalui (i) mendakanmenggunakan kalsium sulfat dan (ii) mendakan menggunakan titik isoelektrik, bila dibandingkan, mengandungi21.6 dan 67.3% protein masing-masing. Kelarutan pekatan pekatan tersebut berkait secara positif dengan pHdalam banjaran nilaipH4-7. Kebolehan membusar berkait rapat dengan percentage ofNlarut (r2 = 038) dan pH(r2 = 0.88) sementara kestabilan busar ada kaitan dengan kehidrofilikan (r2 = 0.98) pekatan. Semua pekatandapat mengurangkan kehilangan berat, kekecutan dan meningkatkan ketegasan sosej ikan yang telah di masak.Peratus kehilangan berat dan kekecutan berkait secara negatif dengan peratus protein larut, pH dan sifat kebolehanmembusar pekatan. Sifat-sifat berfungsi pekatan-pekatan tersebut, bila ditambah kedalam sosej ikan padakandungan 1-2% akan mempengaruhi tekstur produk tersebut. Dalam penilaian deria, sosej ikan yang mengandungiprotein-protein tumbuhan mendapat skoryang lebih tinggi untuk penerimaan keseluruhany walaupun tiada perbezaanbererti dalam skor cita-rasa atau tekstur dan skor yang lebih rendah didapati untuk kesarian produk biladibandingkan dengan hasil kawalan.

ABSTRACTThe physico-chemical and functional properties of mungbean protein concentrate prepared by (i) calcium sulphate

precipitation (MBC-Ca) and (ii) isoelectric point precipitation (MBC-pI) containing 21.6 and 67.3% proteinrespectively, were compared. The solubility of the concentrates was positively correlated with pH within the range of4-7. The foaming ability was closely correlated with percentage of soluble JSf (r2 — 0.98) and pH (r2 = 0.88) whilethe foam stability was correlated with the hydrophilicity (r2 =0.98) of the concentrates. All concentrates were ableto reduce the weight loss, shrinkage and increase the firmness of cooked fish sausages. The weight loss and shrinkagewere negatively correlated with the soluble protein, pH and foaming ability of the concentrates. The functionalproperties of the concentrates, when added at a level of1-2%, influenced the texture of the fish sausages. Inorganoleptic evaluations, fish sausages incorporating the plant proteins scored higher for overall acceptability, eventhough there was no significant difference in flavour or texture and a decrease in juiciness of the product compared tothe control.

INTRODUCTION

Mungbean (Phaseolus aureus) contains about20-27% protein and has an amino acidprofile comparable to soybean (Evans andBandermer 1967; Fan and Sosulski 1974;Thompson et al. 1976). It is an importantprotein source in India, and partiallyreplaces some of the ingredients for babyfoods, snacks and noodles in the Philip-pines, China and Japan (Bhumiratana and

Nondasuta 1972). However, its colour and'beany' flavour limit its use, unless it isdehulled or converted to protein concen-trate. Mungbean protein concentrate(MBC) can be a by-product of mungbeannoodle factories, which only make use of thestarch. MBC may not only improve thenutritional content but also the flavour,texture and appearance of the food.

SUHAILA MOHAMED, JAMILAH BAKAR AND NORHASHIMAH ABD HAM1D

The quality of MBC has been shown tobe comparable to soy protein concentrate(SPC) (Thompson 1975; Bhumiratana1977). Studies on MBC have been limitedto its use as a meat analogue (Narayanaand Narasinga Rao 1982). Its functionalproperties have not been extensively stu-died with regard to its usefulness as astabilizer, thickener, milk substitute, emul-sifier, extender and binder in variousproducts. The functional properties ofMBC merit research for developing its usein food, especially in cereal-based productsbecause of the complimentary amino acidpattern. This work was carried out tocompare selected functional properties ofMBC prepared by two different methodsand the effect of incorporating it in aproduct such as fish sausage.

MATERIALS AND METHODS

Dehulled mungbeans from Thailand ofunknown storage history were obtainedfrom retail shops near the university. Foreignparticles and spoiled beans were removed.Mungbean flour (MBF) was prepared bygrinding the beans in a hammer mill to pass710 nm mesh size sieve (U.S. standardmesh). Commercial grade soya proteinconcentrate from defatted soya flour(Marksaids Malaysia) was used for com-parison (standard) of the functional proper-ties of prepared mung bean concentrate(MBC). Fresh bighead carp {Aristichthysnobilis)) obtained from Salak South, KualaLumpur, were harvested at 6-9 monthsmaturity (about 45-55 cm long and 1.8-2.5kg weight). Unless otherwise stated, allexperiments were carried out at roomtemperature (30 ± 3°C). Preparation ofMBC was-carried out in 6 replicates.

Calcium-precipitated MBC (MBC-Ca)was prepared by extracting MBF withabout 5 times its weight of water andfiltering through a muslin cloth. Theextracts were brought to 90°C, cooled to

85°C, allowed to precipitate for about 30min with 0.4% (w/w) CaSO4 fBDHChemical Ltd, Poole, England), c utri-fuged and dried at 45°C for 24 h beforegrinding and sieving (280nm sieve) (Payu-ma et al. 1985).

Isoelectric point-precipitated MBC(MBC-pI) was precipitated from similaraqueous extracts of MBF by adjusting thepH down to the isoelectric point of themungbean protein ^pH 4.0) with 6N HC1(Analytical grade, Gruppo Montedison,Farmatalia CaHo Erba), centrifuging anddrying as above (Chang and Satterlee1979).

The percentage of soluble nitrogen ofthe protein was determined by the Biuretmethod using bovine serum albumin(Sigma, B2518 lot No33H6780) as thestandard (Layne 1957; Narayana andNarasinga Rao 1982). Emulsifying abilitywas determined by dropping RBD palmolein at 0.2 ml/s into a continuously stirred(Magnetic stirrer) suspension of 2.0 g MBCin 23 ml distilled water (Lin et al. 1974).The end point was when the ammeterneedle (Sanwa, YX-360 TR, Taiwan)suddenly showed a change in reading andthe emulsion separated into two phases.

The foaming ability (Lawhon et al.1972) was the foam volume at 30 s afterhomogenizing 100 ml, 1.0% aqueoussuspension of MBC for 5 min using aKinematica emulsifier (Switzerland) asmeasured using a 250-ml measuring cylin-der. The volume of the foam was monitoredevery 5 min for 120 min. The foam stabilitywas calculated by the formula (2t/50Vm)where Vm =max foam volume (ml), andt = time in min for the foam to collapse toVm/2 (Townsend and Nakai 1983).

Fat-holding capacity (Lin et al. 1974)was determined by measuring the free oilremaining after mixing 0.5g MBC to 5.0 mlof RBD palm olein for 30 s, standing for afurther 30 min, and then centrifuging for 25

70 PERTANIKA J. TROP. AGRIC SCI. VOL. 19 NO. 1, 1996

DIFFERENCES IN FUNCTIONAL PROPERTIES OF MUNGBEAN PROTEIN CONCENTRATE

min at 1750 g. Colour was determinedusing the Hunter-lab colorimeter (modelD25, USA) with a white tile (a = -0.9, b =0.5 and L == 91.25) as the standard. pHwas measured using a pH meter (Jenway,Model PHM64). Moisture was determinedby drying 2.0 g MBC et 105°C to constantweight (AOAC 1980). Crude protein wasdetermined on 0.15 g MBC using themicro-Kjeldahl method where crude pro-tein = N x 6.25 (AOAC 1980).

Preparation of Fish SausageFish sausage was prepared by mixing 50gminced fish flesh, 50g tapioca flour and lgsalt. The proximate composition of'the fishsausage was 16.4% protein, 4.3% fat,76.9% moisture and 1.2% ash. Proteinconcentrate was added to minced fish meatat 0, 1.0, 1.5 and 2.0% level in the presenceof 1 % NaCL They were mixed in aKenwood Chef mixer (model A901) at aspeed of 4 rev/s for 5 min at roomtemperature, then inserted into 35-mmdiameter cellulose casing and tied at theends. After labelling, the sausages were keptfrozen (at -20°C) for 24 h to allow for theformation of intermolecular crosslinks.

Analysis of the Fish SausageFrozen fish sausage was thawed at 30°C for30 min, boiled for 25 min, then air cooledfor 15 min.

The weight loss and shrinkage weredetermined by comparing the averageweight and circumference of six sausagesbefore and after boiling.

The texture of the sausage was empiri-cally determined using an Instron Univer-sal testing machine (Model 1140) with a 5-cm long puncture probe on thawed sausagesamples at a crosshead speed of 50 mm/min,a 5 kg load cell and noting the yield/breaking stress.

The cooked fish sausages were organo-leptically evaluated by 10 trained panellists

for colour, flavour, texture, juiciness andoverall acceptability on a 1-7 hedonic scale(7= extremely like; 1= extremely dislike)(Larmond 1982). The data were analysedusing multiple-range test (Walpole 1982)and regression analysis.

RESULTS AND DISCUSSION

The protein content of both MBCs wasfound to be lower than that of SPC (Table1). The solubility of the protein waspositively correlated (r =0.83) with pHwithin the range studied independently ofhow they were prepared. Therefore, theo-retically the solubility could be adjusted asrequired by bringing the pH away from theisoelectric point of the protein and increas-ing the net charge.

Regression analysis shows that theamounts of moisture in the MBCs andSPC after preparation were positivelycorrelated with the percentage of solublecrude protein of the concentrates (r2 = 0.99),indicating that the hydrophilic nature ofthe soluble protein has some influence onthe moisture content of the concentrates.

Emulsifying Capacity and Fat AbsorptionCapacity

For all concentrates, the values of fatabsorption capacity and emulsifying capa-city followed a similar trend; the highestvalue was seen in MBC-pI and the lowestvalue in MBC-Ca (Table 1). Voutsinas andNakai (1983) showed that a close relation-ship exists between fat binding capacityand surface hydrophobicity, while Katoand Nakai (1980) found a significantcorrelation (P < 0.01) between theemulsifying capacity and the hydrophobi-city of proteins. The present study confirmsthe close relationship between fat-bindingcapacity and emulsion capacity, and thatthey most likely relate to the surfacehydrophobicity of the protein molecules.The result above thus indicates that the

PERTANIKA J. TROP. AGRIC. SCI. VOL. 19 NO. 1, 1996 71

SUHAILA MOHAMED, JAMILAH BAKAR AND NORHASHIMAH ABD HAMID

TABLE 1Physico-chemical properties of calcium sulphate precipitated mungbean protein concentrate [MBC-Ca],

isoelectric point precipitated mungbean protein concentrate [MBC-pI] and soya protein concentrate[SPC]

Sample

crude protein (%)moisture (%)soluble N* (%)PHfat absorption(g oil/g protein)emulsifying capacity(g oil/g protein)foaming ability (ml)0.5 min5.0 min120 minBest fitted lineY =r2 =rate of foamcollapse (ml/min)foam stability(2t/50Vm)t= (min)time to reach Vm/2colour(L) lightness(a) + red, -green(b) + yellow, -blue

MBC-Ca

51.6c±0.610.35b± 10.052.6c±0.75.71b±0.011.9a±0.3

15.2c±0.6

12.07.34.3

6.84-0.016x0.740.02

0.17

50

84.88a±0.05-0.72a±0.0812.77a±0.I9

MBC-pI

67.3b±4.110.18b±0.142.1c±0.54.06c±0.042.4a±0,7

30.2a±10.8

3.93.90.5

3.015-0.023x0.850.02

0.46

45

84.40b±0.03-2.15b±0.0318.05b±0.03

SPC

77.1a±2.212.08a±0.056.3a±0.47.01a±0.0032.3a±0.6

21.7b±0.4

36.036.018.7

36.17-0.145x0.990.15

0.14

125

82.35c±0.03-1.99c±0.0530.38c±0.11

MBF (mungbean flour)

22.3d ±1.6

3.6b±0.36.25

* = pH unadjusted

Means within the same row followed by the same letter are not significantly different (p > 0.05). The standarddeviations were calculated from at least 6 replicates

surface hydrophobicity of MBC-pI is muchgreater than of MBC-Ca. At the isoelectricpoint, the electrostatic charges on therandomly coiled heated protein moleculeinteract, causing exposure of more hydro-phobic side chains (Bigelow 1967) andprotein insolubility.

However, there was poor correlationbetween emulsifying capacity or fat absorp-tion and protein solubility. This agrees withthe work of Wang and Kinsella (1976) whofound no statistical correlation betweenemulsifying capacity and protein solubi-lity, but a high correlation (r2 = 0.8) bet-ween emulsion stability and protein solubi-

lity. The emulsifying properties of proteinsu l t imate ly depend on the hydro -phile:lipophile balance, and do not neces-sarily increase as the proteins become morehydrophobic (Rand 1976). The ability ofproteins to bind lipids is important forapplications such as meat replacers andextenders.

In this study, under the same rates ofblending, oil addition and temperature, thefat absorption capacity and emulsioncapacity were found to be correlated onlywith the protein content in the concentrates(r2 = 0.87 and 0.52 respectively) and notwith pH.

72 PERTANIKA J. TROP. AGRIC. SCI. VOL. 19 NO. I, 1996

DIFFERENCES IN FUNCTIONAL PROPERTIES OF MUNGBEAN PROTEIN CONCENTRATE

Foaming Ability

The foaming ability was very closelycorrelated with the percentage of solubleN (r2 = 0.98), and also with pH of theconcentrate (r2 = 0.88), as Cheftel et al.(1985) and Townsend and Nakai (1983)respectively had also found. The availabil-ity of mixtures of acidic and basic proteins(opposite charges) is thought to be impor-tant for the inter-molecular electrostaticinteractions and strength of the 'skin'around the air bubbles (Hart 1986).

Foaming ability was found not to berelated to emulsion capacity or fat absorp-tion capacity, but surface hydrophobicity,which confirms the work of Townsend andNakai (1983). The presence of salt in thefinal product (as with MBC-Ca and MBC-pl preparation) also reduces foam capacityand stability (Graham and Phillips 1976),but increases emulsion capacity (Wang andKinsella 1976) due to the unfolding of theprotein. The salt content or ionic strengthof the MBCs was not determined in thiswork.

Foam Stability

Rate of foam collapse was found to beclosely correlated (r2 = 0.98) to the water-holding ability (hydrophilicity) of theconcentrates, the soluble nitrogen content(r2 = 0.97) and the foaming ability(r2 = 0.92). Foam stability is positivelycorrelated with emulsion capacity(r2 = 0.75) and negatively correlated withpH (r2 ~ 0.86) and foaming ability(r2 = 0.56). Hydrophobes are thought tocause foam collapse by competing withprotein at the bubble surface, thus disrupt-ing the continuity of the adsorbed proteinfilm (Hart 1986).

Colour of MBC

There is little colour difference betweenMBC-pI and SPC. The colour was found tobe affected by pH, moisture, protein and fat

content and soluble N of the concentrates.

Effect of the MBCs on the Properties of FishSausage

When the MBCs were added, the propertiesof the concentrates affected the properties ofthe fish sausage (Figs. 1 and 2). Thepercentage of protein in the concentrate ispositively correlated with the organoleptictexture score (r2 = 0.75). The increase infirmness with the addition of MBCs waspreferred by the panellists over the control,while addition of SPC was disliked.

The percentage of soluble nitrogen inthe protein concentrates favourably af-fected the sausage weight loss (r2 = 0.78),shrinkage (r2 = 0.71) and Instron-measuredfirmness (r2 = 0.92). The fish sausagesamples which contained protein concen-trates were preferred to the normal fishsausages for flavour and texture.

Increasing the pH of the proteinconcentrates reduced weight loss(r2 = 0.50), shrinkage (r2 = 0.89) and In-stron measured firmness (YZ = 0.85) butdecreased the flavour (r2 = 0.55) andsensory scores for juiciness (r2 = 0.83). Thiscan be attributed to the increased netcharge on the MBCs with increasing pH

control

2% MBF

1.5% MBF

1% MBF

2%

t/

SPC1.5%

1%

\

7NSPC

MBC-Ca15

*4o/

1m1%

1.5% MBC-Ca

2% MBC-Ca

yJ V - 1% MBC-pl

S " ^ n 5o/o MBC_p

\\

2% MBC-pl

SPC

% weight loss % shrinkage firmness/10 Kg

Fig. L Effect of protein concentrate on weight loss,shrinkage and firmness offish sausages

PERTANIKA J. TROP. AGRIC. SCI. VOL. 19 NO. 1, 1996 73

SUHAILA MOHAMED. JAMILAH BAKAR AND NORHASHIMAH ABD HAMID

1% MBC-Cacontrol 7 1.5% MBC-Ca

2% MBC-Ca

1% MBC-Cacontrol -~r-JL 1-5% MBC-Ca

2% MBF

1% MBF

1% MBC-pl 1.5% MBF j

1.5% MBC-pl

2% SPC1.5% SPC

2% MBC-pl

1% SPC

1.5% MBC-pl

2% SPC1.5% SPC

2% MBC-pl1% SPC

colour texture juiciness flavour overall

Fig. 2. Sensory evaluation offish sausages containing plant proteins

above the pi, thus increasing the swellingand the water-binding ability of the MBCs.

The emulsifying capacity and fatabsorption capacity do not have mucheffect on the fish sausage. This functionalproperty may have more influence in meatsausages, which have a greater percentageof fat included in the formulation.

The MBC foaming ability greatlyinfluenced the fish sausage quality, and isnegatively correlated with weight loss(r2 = 0.99), shrinkage (r2 = 0.96), flavour(r2 = 0.99), and texture (r2 = 0.67), butpositively correlated with firmness(r2 = 0.97) and colour (r2 = 0.96). Thefoaming ability is believed to be a measureof the electrostatic interacting capacity ofthe various proteins (Hart 1986) in theconcentrate, and this explains why increasein foaming ability is related to reducedweight loss, shrinkage, texture and flavour.

The foam stabilizing ability of theprotein concentrate is negatively corre-lated with shrinkage (r2 = 0.99) and weightloss (r =0.56), and has been shown earlierto be correlated with the hydrophihcity of

the concentrate.In all experiments the weight loss offish

sausage was reduced by adding proteinconcentrate (Fig. 1). The original sausageweights were better retained with SPC >MBC-Ca > MBC-pl > MBF. Weight loss ismainly due to the reduction in water-holding capacity of the heat-denaturedprotein. Adding the protein concentratesalso increased the firmness of the fishsausages in the fo l lowing o r d e rS P O MBF > MBC-Ca > MBC-pl. Sen-sory evaluation (Fig. 2) showed that thesamples of fish sausage with added plantprotein were preferred (overall acceptabil-ity) to the control sausages. Generally,there was no significant difference intexture and flavour, but a decrease injuiciness was observed in the fish sausageswith added plant protein. Colour scores forthe sausages was higher writh the addition ofSPC. The colour scores for sausages withadded MBC-Ca were insignificantly differ-ent from those of the control, but those withMBF or MBC-pl scored slightly unfavour-ably.

74 PERTANIKA J. TROP. AGRIC. SCI. VOL. 19 NO. I, 1996

DIFFERENCES IN FUNCTIONAL PROPERTIES OF MUNGBEAN PROTEIN CONCENTRATE

CONCLUSION

Incorporation of mungbean protein con-centrate increased the overall acceptabilityoffish sausages by reducing the weight loss,shrinkage and increasing their firmness.

REFERENCESAOAC 1980 Official Methods of Analysis. Washing-

ton D.C. Assoc. of Official Analytical Che-mists.

BHUMIRATANA, A. 1975. High protein low costfood. Tech. paper. Third Asean Sub-committee onProtein. Manila, 1-42.

BHUMIRATANA, A. and A. NONDASUTA, 1972.Report on protein food development project.Bangkok: Nutr. Div. Health Department,Ministry of Public Health, Thailand.

BlGELOW, C.C. 1967. On the average hydro-phobicity of proteins and the relation betweenit and protein structures. Journal of TheoreticalBiology 16: 187-211.

CHANG, K.C. and L.D. SATTERLEE. 1979.Chemical, nutritional and microbiologicalquality of protein concentrate from culleddry beans, Journal of Food Science 44: 1589-1593.

CHEFTEL, J.C., J.L. CUQand D. LORIENT. 1985.Amino acid, peptides and proteins. In FoodChemistry, ed. O. Fennema, 2nd edn. p.245-370. N.Y.: Marcel Dekker.

EVANS, R.S. and S.C. BANDERMER. 1967.Nutritive values of legumes seed protein.Journal of Agriculture and Food Chemistry 15: 439.

FAN, T.V. and F.W. SOSULSKI. 1974. Dispersi-bility and isolation of proteins from legumeflours. Journal of Food Science and Technology 7:256.

GRAHAM, D.E. and M.C. PHILLIPS. 1976. Theconformation of proteins at the air-waterinterface and their role in stabilizing foams.In Foams', ed. RJ . Akers, p. 237-256. N.Y.:Academic Press.

HART, RJ. 1986. New functional ingredients.Food June 35-37.

KATO, A. and S. NAKAI. 1980. Hydrophobicitydetermined by a fluorescent probe methodand its correlation with surface properties ofproteins. Biochemica Biophysica Acta 624: 13-20.

LARWOND, E. 1982. Laboratory methods forsensory evaluation of foods. Technical paper.

Ottawa: Canada Dept. Agric.LAWHON. J.T., CM. CATER and K.F. MATTIL.

1972. A comparative study of whippingpotential of an extract from several oilseedflours. Cereal Science Today 17: 240.

LAYNE, E. 1957. Spectrophotometric and turbidi-metric method for measuring protein. Methodsin Enzymology 3: 447-454.

LIN, M.Y.J., C.S. HUMBAR and F.W. SOSULSKI.1974. Certain functional properties of sun-flower meal products. Journal of Food Science39: 368-370.

NARAYANA, K. and M.S. NARASINGA RAO.1982. Functional properties of raw and heatprocessed winged bean flour. Journal of FoodScience 47: 1534-1538.

PAYUMA, E.M., E.F. FOBIAN, B.S. CORPUZ andL.S. MONGUOIAT. 1985 Mungbean proteinmeat analogue. ASEAN Food Journal 85: 94.

RAND, R.R. 1976. In Food Emulsions, ed. S.Friberg, p.277-294. N.Y.: Marcel Dekker.

THOMPSON, L.U. 1977. Preparation and evalua-tion of mungbean protein isolates. Journal ofFood Science 42: 202-205.

THOMPSON, L.U., L. HUNG, N. WANG, V.RASPER and H. GRADE. 1976. Preparationof mungbean flour and its application inbread making. Canadian Institute of Food Scienceand Technology Journal 9(1): 1-5.

TOWNSEND, A.A. and S. NAKAI. 1983. Relation-ship between hydrophobicity and foamingcharacteristics of food protein. Journal of FoodScience 48: 588-591.

VOUTSINAS, C.P. and S. NAKAI, 1983. A simpleturbidimetric method for determining the fatbinding capacity of proteins. Journal ofAgriculture and Food Chemistry 31: 58-63.

WALPOLE, E.R. 1982. Introduction to Statistics.N.Y.: Mac William Pub. p. 398-402.

WANG, J.C. and J.E. KlNSELLA. 1976. Func-tional properties of novel proteins: Alfafa leafprotein. Journal of Food Science 41: 286-292.

(Received 17 December 1994)

(Acceped 1 April 1996)

PERTANIKA J. TROP. AGRIC. SCI. VOL. 19 NO. 1, 1996 75