Determination of inulin content of chicory roots ( Cichorium intybus L.) cultivated organically in three regions of Rio de Janeiro state.

S. Couri1, F. S. Gomes1 , R. I. Nogueira1 , V. C. Wilberg1, L. M. C. Cabral1, V. V. Silva2, D. L. Almeida3

1

Embrapa Agroindstria de Alimentos: scoury@ctaa.embrapa.br 2 Pesagro-Niteroi 3 Embrapa Agrobiologia

Abstract: Inulin is a mixture of fructose-chains varying from 2 to 60 fructose-units. This carbohydrate is the energy reserve in about 36000 plants, including chicory roots ( Cichorium intybus L.). Inulin was identified as an ingredient which substitutes fat or sugar in the diet. It presents low calorie properties and acts as a bifid factor, which implies on the regulation of the intestinal flora. This carbohydrate is no commercially produced in Brazil. This work had as objective to evaluate the yield and inulin content of chicory cultivated in three different soil and climatic conditions of Rio de Janeiro state (Avelar, Seropdica and Friburgo). Inulin was extracted from fresh and dried chicory roots by means of hot water and precipitated by cooling down. The samples were analyzed by high pressure liquid chromatography (HPLC), using a Aminex HPX-87C (250x4.0mm) column. With regard to the inulin content, the plants cultivated in Friburgo showed 17.17g inulin/100g fresh root. The roots produced in Seropdica and Avelar have, respectively, 16.16 and 15.79g inulina/100g fresh root. These preliminary results are not indicative, however, that Nova Friburgo has the best soil and climatic conditions for the cultivation of C. intybus L. New experiments are being lead to elucidate whether the culture and climate conditions of the studied regions are the determinative factors for the success of chicory root cultivation in Rio de Janeiro state.

Keywords: Inulin, chicory, Cichorium intybus L, fructan, storage carbohydrates

1. Introduction
Chicory (Cichorium intybus L.) is a plant whose tuberous roots store inulin, with a high fructose content (about 94%). It is a reserve carbohydrate, which consists of a long chain made up of 22-60 fructose molecules with a terminal glucose molecule. Inulin is a blend of fructan chains found widely distributed in nature as plant storage carbohydrate. It may be classified as a fructoligosaccharide (FOS), and is present in more than 36,000 plant species, mainly plants of the Asteraceae and Graminae families (Carpita et al., 1989; Van Loo et al., 1995). Chemically, inulin is a polydisperse -(2,1) fructan (Phelps, 1965). The fructose units in this mixture of linear fructose polymers and oligomers are linked to each other by -(2,1) bonds. A glucose molecule typically resides at the end of each fructose chain and is linked by an -(1,2) bond, as in sucrose. The average molecular weight and degree of polymerization of inulin depend upon the source of inulin, the time of harvest and the process of production. Most of the inulin commercially available today is obtained from chicory roots (which contains approximately 150 to 200 mg/g inulin and 80 to 120mg/g oligofructose) by extraction at high temperature followed by an ion exchange process (Dysseleer & Hoffem, 1995). In Europe, many varieties of Cichorium intybus L. (Asteraceae) are cultivated and they are highly appreciated for their bitter taste, due to the presence of large quantities of sesquiterpenes (Peters & Van Amerongen, 1998). Inulins are not only important because of their low calorie properties, but also because of the bifid factor, which implies in the regulation of the intestinal flora. Recently, inulin was identified as an ingredient for fat or sugar

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substitution. Due to these properties, food and pharmaceutical industries have found applications for inulin in the production of functional foods, nutritional composites and medicines. It acts in the organism in a similar way as dietary fibers, contributing to the improvement of the gastrointestinal system conditions (Roberfroid et al., 1993). Limami et al. (1993) studied the influence of the nitrogen fertilization on C. intybus during the cultivation and storage of the roots. The treatments were 0.60g and 120g nitrogen /10m. The root weights during cultivation, harvest and storage at 0.5C were observed. The increase of fructan content in the last periods of growing was associated with the nitrogen increase in the roots. Mehta et al. (1995) evaluated the effect of the spacing and concluded that a 0.30m spacing between plants was better that a 0.20m spacing, while inhibiting competition between plants. Since inulin is not commercially produced in Brazil, this work had as objective to evaluate the yield and inulin content of chicory cultivated in three different soil and climatic conditions of Rio de Janeiro state (cities of Avelar, Seropdica and Nova Friburgo).

2. Materials and methods

2.1. Chicory roots Chicory roots, Chichorium intybus L. var. Fredonia , were obtained from the Embrapa Agrobiologia RJ, Brazil.

2.2. Chicory culture Chicory was cultivated under three different soil and climatic conditions in the Rio de Janeiro state (cities of Avelar, Seropdica and Nova Friburgo), using a 25x50cm or 25x25cm spacing between plants, with or without organic fertilization (1kg manure/m2). After completion of the cultivation period, the plants (Figure 1), composed of outer leaves and roots, were harvested and each part separated, weighed (fresh weight) and immediately stored in the freezer (-20oC). Analysis were performed on both roots and leaves, using 100g of plant material. A total of 20 samples of each part of plant was considered in this work.

2.3. Preparation of chicory extracts Root samples were homogenized with water (1:2 w/v) and heated at 120C for 20 min (1 atm.) in a vertical retort Luferco; the treated plant material was then filtered and subjected to chromatography.

2.4. Extract analysis Sample analysis was performed using a Waters high performance liquid chromatograph under the following conditions: column - Aminex HPX-87C (300x7.8 mm, i.d); detector - refractive index detector, Waters model 2414; eluent: water; flow rate: 0.3ml/min; injected volume: 20.0l; column temperature: 80oC; detector temperature: 40oC (Nogueira, 2002). For the inulin quantitation, a commercial standard (Fluka-BioChemika 57614) was used.

2.5. Composition of chicory roots Protein, moisture, fiber, ash and oil contents in the raw material (roots) were determinated according to AOAC standard methods and the carbohydrate content calculated by difference.

Fig. 1. Chicory plants (Cichorium intybus L.) showing outer leaves and tuberous roots.

3. Results
Except the Seropdica 1 and Seropedica 6 samples, the Cichorium intybus L, cultivated in the 3 regions of Rio de Janeiro presented inulin concentration around 15g/100g of fresh roots. These results are compatible with data found in the literature. Baert (1997) studied three cultivars varying the time of plantation and harvest and achieved an inulin yield between 15.2% and 16.5% in the harvests in September. Amaducci & Pritoni (1998) also evaluated the effect of time of harvest for five cultivars and achieved a inulin yield in the roots around 15%. Some differences are found when considering the culture conditions shown in Table 1. In Avelar, the best culture conditions were spacing of 25cmx50cm and organic fertilization, with a root productivity of 2.67ton/ha. In Seropdica, the best conditions were spacing of 25cmx25cm without organic fertilization, with a root productivity of 3.57 ton/ha. In Friburgo, 3.62ton/ha of roots were obtained, using a 25cmx50cm spacing and without organic fertilization. These preliminary results indicate that Seropdica and Friburgo regions were more adequate for chicory plantation than the Avelar region. However, no final conclusions could be drawn with the obtained spacing and fertilization data. New experiments are being carried out to elucidate the best culture and climate conditions for the chicory root development, aiming at inulin obtention in the Rio de Janeiro state. Considering the value of 15 g inulin/100g of fresh chicory roots (Table 1) as an average of the analyzed samples, the content of other carbohydrates of chicory roots varied from 18 to 23% as shown in Table 2. The Seropdica 6 sample presented the lowest inulin concentration in the roots. So, other root storage carbohydrates represent about 11.70g/100g in this sample. Evaluating the root protein content, the variation was from 0.51 to 1g/100g of fresh chicory. The small differences in composition of the samples have no influence on root quality , considering that the main objective of this work was to achieve high concentration of inulin in the roots.

Table 1. Chicory roots production and inulin yield.

Sample

Spacing (cm)

Avelar 1 Avelar 2 Avelar 3 Avelar 4 Seropdica 1 Seropdica 2 Seropdica 3 Seropdica 4 Seropdica 5 Seropdica 6 Seropdica 7 Seropdica 8 Friburgo 1 Friburgo 2

25x50 25x50 25x25 25x25 25x50 25x50 25x25 25x25 25x50 25x50 25x25 25x25 25x50 25x25

Organic fertilization (kg manure/m2) 0.0 1.0 0.0 1.0 1.0 0.0 1.0 0.0 0.0 1.0 1.0 0.0 1.0 0.0

Yield whole plant (ton/h)

Root productivity (ton/ha) 2.22 2.67 1.83 2.08 1.72 2.15 2.46 3.57 2.25 2.11 2.83 2.25 3.62 2.90

g inulin/100g fresh chicory roots 12.80 13.48 13.66 15.79 11.00 14.69 15.04 15.83 14.17 7.26 15.19 16.16 17.17 12.24

4.67 5.50 4.83 4.58 4.31 4.70 6.60 9.45 5.79 5.25 7.47 7.50 6.00 4.50

Table 2. Composition of chicory roots.

Sample Avelar 1 Avelar 2 Avelar 3 Avelar 4 Seropdica 1 Seropdica 2 Seropdica 3 Seropdica 4 Seropdica 5 Seropdica 6 Seropdica 7 Seropdica 8 Friburgo 1 Friburgo 2

Ash (g/100g) 1.08 1.12 1.06 1.08 1.14 1.14 1.06 1.14 0.99 1.07 1.02 0.99 0.99 1.43

Moisture (g/100g) 75.74 76.89 76.97 76.43 77.74 77.28 78.17 77.09 79.50 78.88 78.33 77.30 74.40 75.84

Protein (g/100g) 0.81 1.10 0.53 0.59 0.69 0.56 0.56 0.50 0.81 0.87 0.62 0.62 1.0 0.94

Oil (g/100g) 0.21 0.36 0.15 0.12 0.62 0.60 0.39 0.48 0.20 0.23 0.22 0.31 0.53 0.73

Carbohydrate (g/100g) 22.16 20.53 21.29 21.78 19.81 20.42 19.82 20.79 18.50 18.95 19.81 20.78 23.08 21.06

4. Conclusions
The preliminary results are only indicative that the region of Friburgo has the best soil and climatic conditions for the cultivation of C. intybus L. New experiments are being carried out to verify whether the culture and climate conditions of the studied regions are the determinant factors for the success of chicory root cultivation in the Rio de Janeiro state.

References
Association of Official Analytical Chemists AOAC, Official methods of analysis of Association Of official Analytical Chemists. Washington DC, 1995. Carpita, N. C., Kanabus, J., and Housley, T. L. 1989. Linkage structure of fructans and fructan oligomers from Triticum aestivum and Festuca arundinacea leaves. J. Plant Physiol. 134: 162168.

Dysseleer, P, & Hoffem, D. I. ( 1995). An alternative dietary fibre. Properties and quantitative analysis. European Journal of Clinical Nutrition, 49 ((3S)), S 145-S152. Glyn Maria Figueira , Kil Jin Park, Fernando Pedro Reis Brod, Sylvio Luis Honorio. Evaluation of desorption isotherms, drying rates and inulin concentration of chicory roots (Cichorium intybus L.) with and without enzymatic inactivation.Journal of Food Engineering 63 (2004) 273-380. Nogueira, R. I. (2002). Processo de obteno de inulina de chicria (Cichorium intybus) em p. Doctorate Thesis. School of Agricultural Engineering, University of Campinas, Brazil. Peters, A.M., & Van Amerongen, A. (1998). Relationship between levels of sesquiterpene lactones in chicory and sensory evaluation. Journal of the American Society for Horticultural Science, 123, 326329. Phelps, C. 1965. The physical properties of inulin solutions. Biochem. J. 45: 4147 Roberfroid, M., Gibson, G. R., & Delzenne, N. (1993). The biochemistry of oligofructose, a nondigestible fiber: an approach to calculate its caloric value. Nutrition Reviews, 51(5), 137146. Van Loo, J., Coussement, P., Leenheer, L. D., Hoebregs, H., and Smits, G. 1995. On the presence of inulin and oligofructose as natural ingredients in the western diet. Crit. Rev. Food Sci. Nutr. 35: 525552.

Acknowledgments
The authors would like to thanks FAPERJ- Fundao de Amparo Pesquisa do Estado do Rio de Janeiro, and CNPq-Conselho Nacional de Desenvolvimento Cientfico e Tecnolgico, Brasil for research financial support and scholarships.