Starch Update 2011: The 6 th International Conference on Starch Technology

P-STARCH-4

Functionality benchmarking of underutilized starches with cassava starch

Sirithorn Lertphanich 1, Rungtiva Wansuksri 2, Thierry Tran 3, Guillaume Da 4, Luong Hong Nga 5, Kuakoon Piyachomkwan 2 and Klanarong Sriroth 1

1Department of biotechnology, Faculty of Agro-Industry, Kasetsart University, Bangkok, Thailand 2Cassava and Starch Technology Research Unit, National Center for Genetic Engineering and Biotechnology (BIOTEC), Bangkok, Thailand 3Centre de Coopération Internationale en Recherche Agronomique Pour le Développement (CIRAD), Dpt. Persyst, UMR Qualisud, Montpellier, France 4 CERTES, Université Paris-Est Créteil, 61 avenue du Général de Gaulle, 94000 Créteil, France 5 Hanoi University of Science and Technology (HUST), IBFT, Hanoi, Vietnam

Abstract Plant organs represent a highly diverse source of reserve starch but many crops remain underutilized for agro-industries. In this study, structural and functional properties of starches from various storage organs including from tubers, i.e.yam bean ( Pachyrhizus erosus ), from corms, i.e.taro ( Colocasia esculenta ) and ensete ( Enseteventricosum ), from fruits, i.e. water caltrop ( Trapa natans ) and from grains (legumes), i.e. chickpea (Cicer arietinum ) and mungbean ( Vigna radiata ) were characterized and compared with cassava starch. All extracted starches were pure (protein and ash < 0.25% dwb). Taro starch had the smallest granules (2 µm) while ensete granules were the largest (42 µm). Only ensete starch had the B-type polymorph, while mungbean and chickpea starches were C-type and the rest were A- type. Yam bean, water caltrop and ensete starches had amylose contents of 14.15, 19.20 and 20.85%, respectively, similar to cassava starch (17.44%), while taro starch was lower (7.91%) and mungbean and chickpea starches were higher (28.47% and 35.59%. With regard to functional properties, different gelatinization, pasting and rheological properties were observed. Based on peak viscosity as determined by Rapid Visco Analyzer (9.2% starch dsb), three groups of starches were classified; high (cassava and mungbean; 319 and 373 RVU), medium (yam bean, water caltrop and ensete with 222-281 RVU) and low (taro and chickpea with 197 and 176 RVU, respectively). Interestingly, only water caltrop, mungbean and chickpea had a positive value of setback from peak, implying high tendency of gelation. The dynamic rheological analysis also revealed high storage modulus (G ′) values upon cooling for those three starches, while cassava, yam bean and taro had low G ′ values. Taro had the highest susceptibility to α-amylase hydrolysis, while ensete starch granules had the lowest. These results demonstrate various starches with diverse properties, comparatively to cassava starch and can potentially find value-added uses as food ingredients.

Keywords: Cassava, ensete, water caltrop, chickpea, yam bean, taro, mungbean, paste, rheology

1. Introduction Starch is the second most abundant polysaccharide after cellulose as an energy reserve in many photosynthesizing plants. It is found in different storage organs including seeds (corn, wheat, rice, barley, sorghum), tubers (potato, yam), roots (cassava), fruits (mango, banana), stems (sago palm), rhizomes (canna), corms (taro).These are employed as an important staple food in many regions and are also used for starch production. Extracted starches are extensively employed in a wide range of food and non-food application, functioning as thickeners, binders, emulsifiers, texture modifiers, gelling agent, film forming agent and so

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on. Nowadays, major commercial starches are produced from seeds, i.e. corn, wheat and rice, from tubers, i.e. potato and from roots, i.e. cassava. In tropical areas where high plant biodiversity is recognized, there are still many diversified starch-reserved plants being underutilized. Starches from different botanical origins are likely to have diversified structural and physico-chemical properties. In this study, properties of starches from various underutilized crops grown in tropical regions were characterized and compared with cassava starch.

2. Materials and Methods 2.1. Materials Cassava (Manihot esculenta ), Mungbean (Vigna radiat a),Taro (Colocasia esculenta ), Yam bean (Pachyrhizus erosus ), Chickpea(Cicer arietinum ) were collected from local markets in Thailand. Water Caltrop (Trapa natans ) was taken from Vietnam. Ensete(Enseteventricosum ) was obtained from a local market in Ethiopia.

2.2 Methods Starch extraction and composition analysis Starch was extracted by grinding peeled samples with water and passing through a 170-mesh sieve. Starch slurry was then collected and settled. After starch sedimentation, water was decanted and starch cake was rewashed twice before drying. Starch purity was determined by an enzyme method (AACC, 1995). If the purity was less than 90%, the sample was further washed with sodium hydroxide solution. The content of protein, fat, fiber and ash were also quantified according to the AOAC methods (AOAC, 1990).

Structural properties: Granule morphology and amylose content Granule morphology of starch samples was observed under JEOL scanning electron microscopy (JSM-5310, England) at 10-KV acceleration. The X-ray patterns of starches were obtained with Cu-K∝ radiation using a diffractometer (Joel X-ray diffractometer JDX-3080, Joel, Japan), scanning from 5 ° to 40 ° 2 θ at 0.02 ° 2 θ/min. Iodine affinity measurement by an automatic amperometric Titrator (835 Titrando,Metrohm, Herisau, Switzerland) was performed to quantify the amylose content, according to the method of Takeda & Hizukuri (1987).

Viscosity properties Viscosity properties of the starch suspension (9.2 % dwb) were evaluated using a Rapid Visco Analyzer (RVA4, Newport Scientific, Australia). The starch suspentsion was heated from 50 to 95 °C at a rate of 9 °C/min and held at 95 °C for 3 min . After that, starch paste was cooled down to 50 °C at the same rate. The viscosity of starch paste, including peak viscosity (P), viscosity at 95 °C (H), viscosity at 50 °C (C) and gelatinization temperature were recorded. The breakdown (P-H) and setback from peak (C-P) were also reported.

Rheological measurements Rheological properties of the starch suspensions (20%w/w) were determined on a rotational Physica MCR 300 rheometer (Physica Messtechnik GmbH, Stuttgart, Germany) with a plate and plate geometry sensor (50 mm diameter, and 1mm gap). A thin layer of light paraffin oil was added to prevent evaporative loss. Temperature ramp sweep test of the samples was performed from 25 to 95 °C, and from 95 to 25 °C at a rate of 1.5 °C/min. Parameters including storage modulus (G ′), loss modulus (G ′′) and loss tangent (tan δ = G ′/G ′′) were monitored at frequency of 1 Hz and a strain of 0.5 % (thelinear viscoelasticity domain).

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In vitro digestibility In vitro starch digestibility was analyzed according to the method of Miao et al. (2009). The starch sample (200 mg) suspended and prewarmed in phosphate buffer (15 ml, 0.2 mol/l, pH 5.2,37 oC) were incubated with porcine pancreatic α-amylase (290 U/ml) and amyloglucosidase (15 U/ml)with shaking (150 rpm). Aliquots of hydrolysed solution (0.5 ml) were collected at various reaction times. The glucose content of centrifuged hydrolyzate was determined using the glucose oxidase-peroxidase assay kit (Megazyme, Wicklow, Ireland).

3. Results and Discussion 3.1 Starch composition analysis Extracted starches prepared in this study were quite pure as indicated by very low quantities of impurities, i.e. protein, lipid and ash (in total not greater than 0.5% dwb). The starch contents by enzymatic assay of all extracted samples were greater than 95% dwb.

3.2 Granule morphology and amylose content Starch granule morphology was discrete and species-specific (Table1). Taro starch granules were polyglonal with the smallestsize of 2 µm, followed by yam bean starch with the spherical granules of 8.2 µm in average. Among all starches, ensete starch had the largestangular granules of 42 µm. Granule morphology of mungbean and chickpea starches was quite similar in shape, i.e. oval and round and size (17.7 and 21.0 µm, respectively). The X-ray diffraction displayed an A-type polymorph for cassava, taro, yam bean and water caltrop while ensete starch possessed a B-type crystal polymorph. Similar to typical pea starches, both mungbean and chickpea starches were C-type. The amylose contents of extracted starches were different, ranging from 7.9% in taro starch to 35.6%for chickpea starch (Table 1).

3.3 Pasting properties During heating, insoluble starch granules can absorb water, swell to much larger sizes with some granule disruption and glucan leaching. Accordingly, the starch suspension becomes to paste with changes its flowability. The abrupt increase in paste viscosity of starches from different sources occurred at different temperatures and reached to different maximum viscosity (Table 2). Paste viscosity of ensete starch having the largest granules was developed at the lowest temperature (69.80 °C) while taro starch with the smallest granules had rapid viscosity change at a very high temperature (82.35 °C) with low peak viscosity (197 RVU). The highest peak viscosity of cooked paste was observed in mungbean starch (373 RVU), followed by cassava starch (319 RVU) with high breakdown values, indicating less resistance to shear thinning upon continuous cooking. Interestingly, water caltrop, mungbean and chickpea starches exhibited the positive setback values, i.e. the difference between final and peak viscosity, suggesting a high tendency of chain reassociation, in particular the linear amylose chains, upon paste cooling (Gebre-Mariam, 1996).

3.4 Rheological measurements The structural transitions associated with phase change in starch systems are reflected by changes in the rheological profiles. At early stages of heating, starch granules become swollen and close-packed in accordance with amylose leaching, causing a sol to gel formation and a sudden rise of storage modulus(G′). With further heating, G ′ decrease as caused by a disruption of gel structure (Hsu et al., 2000). The temperature at which rapid G ′ change (T G′) occurred and the maximum G ′ during heating of different starches were varied. Chickpea

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starch gave the highest peak G ′ (13,700 Pa), while taro starch showed lowest values (G ′ = 112 Pa).The loss modulus (G ′′) exhibited similar pattern to that for G ′ during heating (data not shown).Upon cooling, G ′ increased again, suggesting some phase transition was still undertaken (Figure 2).Chickpea, mungbean and water caltrop starches demonstrated relatively steep increase, forming a sigmoidal pattern of modulus development. The highest value was in chickpea. This is in accordance with thehigh tendency of these starch pastes to retrograde, which in turnis due to their very high amylose content (Singh et al., 2007). In contrast, cassava, yam bean and taro had low storage modulus (G ′) values.The tan δ during cooling decreased which might be due to retrogradation of leached components and interaction between molecules remaining inside the granule, reinforcing the gel structure during cooling (Hsu et al., 2000). Consequently, the tan δof starch pastes during coolingcouldto be an evidence of gel formation ability (Table 3). Starches with low tan δ values (water caltrop, ensete, mungbean and chickpea) seemed to retrograde and have gel-like character at a greater extent than starches withhigh tan δ values (cassava, yam bean and taro). Difference in phase transition and relating rheological properties is likely caused by difference in starch structure including amylose/amylopectin ratio, amylopectin organization and granule architecture (Klucinec and Thompson, 2002).

3.5 Digestibility studies The hydrolysis curves of granular starches are shown in Figure 3. The degrees of starch hydrolysis of different starches were varied significantly. Taro had the highest (35.6%) while ensete starch showed the lowest value (2.3%). This could be an effect of granule size. The smaller granules (taro starch) possessed greater surface areas for enzyme accessibility. Yet, other granular structure, e.g. surface pores, interior channels and surface charges can be influenced starch granule hydrolysis. At 180 min of incubation, the degree of hydrolysis of all starches were ranked in an order from low to high as taro (35.6%) > yam bean (16.1%) >mungbean (12.0%) > chickpea (11.6%) > cassava (8.0%) > water caltrop (6.7%) > ensete (2.3%) starches.

4. Conclusion Structural and physico-chemical properties of some underutilized starches were elucidated and were different from cassava starch. Some starches like mungbean, chickpea, water caltrop and water caltrop demonstrated better gelling ability than cassava starch. Instead of introducing chemical modification to cassava starch, a composite blend of cassava starch with those starches can lead to starches with divergent properties for more applications.

Acknowledgements This work is partially supported by the Center for Agricultural Biotechnology under the Higher Education Development Project, Commission on Higher Education, the Ministry of Education as well as by the Agence Universitaire pour la Francophonie (AUF).

References American Association of Cereal Chemists (AACC). 1995. Approved Methods of the American Association of Cereal Chemists, St. Paul, Minnesota. Association of Official Analytical Chemists (AOAC). 1990. Official Methods of Analysis. 15 th ed. The Association of Official Analytical Chemists, Virginia. Gebre-Mariam, T. & Schmidt, P.C.1996. Isolation and physicochemical properties of ensete starch. Starch/starke, 48: 208-214. Hsu, S., Lu, S., & Huang, C. 2000. Viscoelastic changes of rice starch suspensionsduring gelatinization. Journal of Food Science, 65: 215–220.

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Klucinec, J.D., Thompso, d.B. 2002. Amylopectin structure and the amylose to amylopection retio both influence the nature of starch gels from mixtures of amylopection and amylose. Cereal chemistry, 79: 24-35 Miao, M., Zhang, T., Jiang, B. 2009. Characterizations of kabuli and desi chickpea starches cultivated in China. Food Chemistry, 113:1025-1032. Singh, J., McCarthy, O. J., Singh, H., Moughan, P. J., & Kaur, L. 2007. Morphological, thermal and rheological characterization of starch isolated from New Zealand Kamo Kamo fruit – A novel source. Carbohydrate Polymers, 67: 233–244. Takeda,Y. and S. Hizukuri. 1987. Structures of rice amylopectins with low and high affinities for iodine. Carbohydrate research, 168: 79-88.

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Table 2 Pasting properties of various starches as determined by a Rapid Visco Analyzer.

Peak Viscosity Viscosity Setback Pasting Viscosity at 95 oC at 50 oC Breakdown from peak Starch sources Temp ( oC) (P, RVU) (H, RVU) ( C, RVU) (P-H, RVU) (C-P, RVU) Cassava 72.28±0.32 319±0 143±1 193±1 176±1 -126±2 Taro 82.35±0.28 197±4 117±0 177±1 80±4 -22±5 Yam bean 77.08±0.04 281±3 133±2 191±3 148±2 -90±0 Water caltrop 86.80±0.49 222±3 168±2 259±2 54±2 37±0 Ensete 69.80±0.35 277±2 138±2 193±1 139±1 -84±3 Mungbean 77.63±0.25 373±7 272±2 535±3 101±5 162±3 Chickpea 71.28±0.32 176±0 151±1 290±3 25±1 114±3

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0 0 10 20 30 40 50 60 70 80 90 100 Temperature(ºC) Cassava Taro Yambean Water caltrop Ensete Mungbean Chickpea

Figure 1 Storage modulus (G ′) during heating from 25ºC to 95ºC for various starches.

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Cassava Taro Yam bean Water caltrop Ensete Mungbean Chickpea

Figure 2 Storage modulus (G ′) during cooling from 95ºC to 25ºC for various starches.

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Table 3 Dynamic rheological characteristics of various starches during heating and cooling.

**** *** Cooling * ** Heating TG′ Peak G′ o Starch sources ( C) (Pa) G′ (Pa) tan δ G′ (Pa) tan δ Cassava 61.62±0.56 2329±26 259±40 0.24±0.01 410±45 0.23±0.01 Taro 67.51±0.00 107±6 104±1 0.41±0.00 241±0 0.31±0.01 Yam bean 65.18±1.15 1971±81 504±76 0.20±0.00 727±47 0.14±0.01 Water caltrop 74.98±0.56 2513±18 2001±49 0.09±0.01 18615±346 0.04±0.00 Ensete 60.43±0.00 5091±158 1745±72 0.04±0.00 6537±378 0.03±0.01 Mungbean 68.30±2.23 10143±159 3702±513 0.08±0.01 17195±648 0.03±0.00 Chickpea 57.68±0.56 13083±833 6112±362 0.08±0.02 24489±733 0.01±0.00

∗ Temperature at which rapid G′ increase occurs **Maximum G′ value during heating ***All rheological parameter values were obtained at 95ºC **** All rheological parameter values were obtained at 25 °

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35 Cassava

30 Taro

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15 Totalstarch hydrolysis (%) Ensete 10

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Figure 3 Degree of hydrolysis of various starches.

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