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Effect of Maturity Stage on Protein Fractionation, In Vitro Protein Digestibility and Anti-nutrition Factors in Pineapple (Ananas comosis) Fruit Grown

Effect of Maturity Stage on Protein Fractionation, In Vitro Protein Digestibility and Anti-nutrition Factors in Pineapple (Ananas comosis) Fruit Grown in Southern Sudan

Murwan K. SabahelKhier and Saifeldin A. Hussain

Department of Biochemistry, School of Biotechnology, Faculty of Science and Technology, Al Neelain University, Sudan.

E-mail: murwansabahelkhier@yahoo.com

Abstract: This investigation showed that albumin, globulin and protein content increases with increasing the days of maturity stage while glotulin and non protein nitrogen content decreases with increasing the days of maturity stage. The prolamin remained constant during the days of maturity stage. In-vitro protein digestibility improved with decreased of tannin and phytic acid content due to increasing the days of maturity stage because tannin and phytic acid inhibit the activity of pepsin enzymes.

Keywords: Pineapple, Digestibility, Ant-nutritional, Maturity and Vitamins

1.0 Introduction

The importance of pineapple fruit lies in their nutritive value. Fruit of pineapple reached the maturity stage at 105 days after flowering (Guerra and Livera, 1999).

It is rich in protein, fat, fiber, vitamins, mineral salts and water (Atif and Hagag, 1995). The net protein utilization is referred to the percentage of ingestion and utilizing protein for growth and maintenance (Osborne, et al., 1978).It is considered as supplement to daily food (Pandey and Ajanta, 1993). The physical, chemical and sensorial characters of pineapple showed significant difference at the several of maturity stage (Guerra and Livera, 1999). Pineapple contained 25-50 mg/g ellagic acid (Amakura et al., 2000).The juice of pineapple fruit contained five proteolytic enzymes collectively known as bromelain (William, 2000). Bromelain is natural blood thinner because it prevents excessive blood platelet stickiness. It also endues the thickness of mucus, which may benefit the patient with asthma or chronic bronchitis. In addition, bromelain in combination with trypsin may enhance the effect of antibiotic in people with urinary tract infection (Manhart et al., 2002). Bromelain has some remarkable characters such as its ability to reduce inflammation pain and swelling, speed the heeling of injuries trauma, surgery, sinusitis or arthritis (Mackay and Miller, 2003). The pineapple fruit contained

tannin, phenolic acid and Flavonoids (William, 1991).

Objectives of this investigation are study effect of maturity stage (75, 90 and 105

days after flowering) on the protein fractionation, in vitro protein digestibility and antinutrition factors (Tannin, phytic acid)of pineapple fruit..

2.0 Material and methods

2.1 Collection and preparation of sample: The samples were collected from Southern Sudan (Yambio Research Station, Agriculture Research Organization, Ministry of Agriculture, Sudan) basis on maturity stage. The preparation of samples was carried according to the method described by AOAC (1984).

2.2 Protein fractionation

The sequential extraction of protein was carried out according to Mendel and Osborne method (1924). It is basis on solubility of protein in different solvents. Water soluble protein (Albumins), salt soluble protein (Globulins), alcohol soluble protein (Glotulin), alkali soluble protein (Prolamin) and residual proteins (None – protein nitrogen).The residues remaining after those successive extractions with four solvents were determined by semi micro-Kjeldhal method according to AOCA (1990).Percent protein extracted was calculated to total amount of protein in the samples extracted such as follows:

Soluble protein (total) % = T X N X TV X 14 X 6.25 X 100

1000 X A

Protein solubility % = Soluble protein X 100

Total protein

Where:

T = Titer reading (ml of HCl), N = Normality of the HCl (0.02N), TV =Total Volume of the aliquot extracted (100ml), A = Number of (ml) of sample extracted (2.0g), 14 = each ml of HCl is equivalent to 14 mg. Nitrogen, 1000 = Number of mg in one gram and 6.25 = conversion factor from nitrogen into protein %.

2.3 Anti-nutrition factors

2.3.1 Tannin content

Quantitative estimation of tannins for each sample was carried out by using modified vanillin- HCl methanol method as described by Price and Butler (1987).

There is no useful standards curve for tannin in food, but the tanninic acid was used for preparation the standard curve of tannic acid. The standard curve of tannic acid was prepared according to AOAC (1990) for measurement the concentration of tannin in our samples (plotting the concentration of tanninic acid (mg) against the corresponding reading of Spectrophotometer in Absorbance).

2.3.2 Phytic acid content

The phytic acid content was determined according to the method described by Wheeler and Ferrel (1971). Preparation of standard curve for phytic acid was done as follows: standard curve of different Fe (NO3) 3 concentrations was plotted against the corresponding of Spectrophotometer to calculated the ferric iron conc. The phytate phosphorus was calculated from the concentration of ferric iron assuming 4:6 irons: phosphorus molar ratio.

2.4 In vitro protein digestibility: It was carried out according to Maliwal method (1981) in the manner described by Monjula and John (1991) with minor modification. A know weight of the sample containing 16 mg nitrogen was taken in the triplicates and digested with one mg pepsin in 15 ml of 0.1 M HCL at 37 oC for two hours. The reaction was stopped by the addition of 15 ml (10%) trichoroacetic acid. The mixture was filtrated quantatively through filter paper (Whatman No.1). Trichoroacetic acid soluble fraction was assayed for the nitrogen by semi-micro Kjeldhal method.

Protein digestibility % = N2 in supernat – N2 in pepsin X 100

N2 in sample

2.5 Statistical analysis:

Three separate sub samples from each origin sample were taken and analyzed. Then mean values were averaged. Data were assessed by analysis of variance (ANOVA)  as described by Gomez and Gomez (1984).

3.0 Results and Discussion

3.1 Protein fractionation

Table I indicated that albumin content of 105, 90 and 75 days maturity of pineapple is 4.4, 3.2 and 3.0 %, respectively. The results revealed that albumin content increases with increasing period of maturity. Globulin content of 105, 90 and 75 days maturity of pineapple is 1.2 and 0.6, 0.4 %, respectively. The finding indicated that globulin content of fruit increases with increasing the period of maturity. The Prolamin content of 105, 90 and 75 days maturity of pineapple is remained constant (0.7 %) during the period of maturity .Those results revealed that the Prolamin of pineapple not effected by period of maturity. The Glotulin content of 105, 90 and 75 days maturity of fruit is 0.9, 1.0 and 1.8%, respectively. The findings revealed that Glotulin content decreases with increased the period of maturity. Non Protein N2 of 105, 90 and 75 days maturity of fruit is 92.9, 94.6 and 94.2 %, respectively. All the above results are supported by the findings that reported by Amukura et al, (2000).

3.2 In vitro protein digestibility and Anti-nutrition factors

Table 2 illustrated that protein content of 105, 90 and 75 days maturity is 4.11, 3.75 and 3.70 %, respectively. The findings are high than those given by Ahmed (2001).

1. The protein digestibility of 105, 90 and 75 days maturity is 39.4, 36.2 and 32.2 %, respectively. Those results are lower than protein digestibility of sorghum (46 %), and rice (66%), maize (73 %) and wheat (81 %) that given by MacLean (1981). This finding indicated that protein digestibility of pineapple fruit is extremely poor compared with sorghum, rice, maize and wheat. Tannin content of 105, 90 and 75 days maturity is 4.5, 22.2 and 20.2 %, respectively. The results revealed that high tannin content reduced the digestibility of the protein because tannin acts as anti-enzymatic activity. In addition, tannin reacts with protein to form insoluble complex compound. The phytic acid of 105, 90 and 75 days maturity is 0.40, 0.20 and 0.15 mg/g, respectively. Phytic acid interacts with protein forming complex compound and reduce the bioavailability of protein and inhibits the action of pepsin, trypsin and ? - amylase. These results explain the poor of protein digestibility in the pineapple fruit.

References

AOAC (1984).Official Method of Analysis, 14th edition, Published by AOAC Inc.1111 North 19th street, Suite 210 Arlington, Virginia 22209.

AOAC (1990). Official Method of Analysis, 14th edition, Association of Official and Analytical Chemists. Washington, D.C

Ahmed E., (2001). The rare fruit 1st ed., Al Dar Arabia for publication and distribution.

Amukura Y., Okada M., Tsuji S., and Tonogai Y. (2000). Determination of ellagic acid of fresh and processed fruit of pineapple by HPLC. Journal of Food Hygiene Society of Japan 41(3):206 – 211.

Atif M.I.and Hagag N.M.(1995). Ever green fruit, plantation, protection and production 1st Ed. By Association of Almaarifa, Pp 195.

Gomez T.P. and Gomez A.A. (1984). Statistical Procedure for Agriculture Research John Willy and Sons Inc. New York, U.S.A.

Guerra N.B.and Livera A.V.S.(1999). Correlation between the sensorial profile, physical and chemical analysis of pineapple C.V.Perola Revista – Brassilera – fruticulture, abstract 21(1): 32 – 35.

MacLean W.C., Lopez G., de Romana Placko R.P. and Graham G.G. (1981). Protein quality and digestibility of sorghum pre- school Children: Balance studies and plasma free amino acids. J.Nutr. (1):1928 -1936.

Maliwal B.P. (1981). In vitro method to assess the nutritive value of leaf concentrate, J. Agric. Food Chem.31: 315 – 319.

Manjula S. and John E. (1991). Biochemical changes and in vitro protein digestibility of the endosperm of germinate Dolichos lablab. J.Sci. Food Agric., 55:529 – 539.

Manhart N., Akomeak R., Bergmeister H., Spittler A., Ploner M. and Roth E. (2002). Administration of proteolytic enzyme bromelain and trypsin diminish the number of CD4+ cell and the interferon – gamma response in Payer's patches and spleen in endotoxemic balb/c. Cell immunol., 215 (2):113 -119.

MacKay D. and Miller A.I.(2003).Nutritional support  for wound heeling .Altern Med Rev., 8(4):359 -377.

Mendel L.B. and Osborne T.B.(1924). Nutritional properties of protein of maize kernel. J.Biol.Chem.18:1 – 4.

Osborne O.P. and Pvogt G.A. (1987). The analysis of nutrients in food 1st ed. Academic Press, London, New York.

Pandey S.N. and Ajata. (1993). Plant Anatomy Economic, volume 3, Department of Botany DAV College. University of Kanpur, Vikas Publishing House P.V.T.Ltd.

Price M.I.and Butler L.G. (1987). A critical evaluation of the vanillin reactions as an assay for tannin in sorghum rain.J.Agric. Food Chem.26 (5): 1214 – 1218.

Wheeler E.I. and Ferrel R.E. (1971). A method for phytic acid determination in wheat fractions cereal Chem.48:312 -320.

William H. (1991). Evergreen orchards, California University, 2nd ed. Translated by Ghazi Ibrahim and Abdallaal Higazi, Al Dar Arabbia for publication and distribution Pp 483.

William C.F.(2000). Trease and even Pharmacognosy. Formerly Reader in phytochemistry, University of Nortingham, Nortingham UK 15th ed Saundes Edinburgh London – New York Oxford Philaddeiphia, St – Lous Sudneytoranto.

Table 1: Protein fractionation of 105, 90 and 75 days maturity of pineapple fruit, grown in Southern Sudan, 2007.

Sample

Albumin

Globulin

Prolamin

Glotulin

Non Protein N2

105 days Maturity

4.4b

(±0.03)

1.2a

(±0.6)

0.7a

(±0.13)

0.9a

(±0.03)

92.9b

(±0.01)

90 days Maturity

3.2a

(±0.09)

0.6a

(±0.05)

0.7a

(±0.13)

1.0a

(±0.12)

94.6a

(±0.01)

75 days Maturity

3.0a

(±0.09)

0.4a

(±0.05)

0.7a

(±0.13)

1.8a

(±0.12)

94.2a

(±0.01)

• Mean values of same letters within the column are significantly difference at

(P? 0.05).

Table 2: In Vitro protein digestibility and Anti-nutrition factors of 105, 90 and 75 days maturity of pineapple fruit, grown in Southern Sudan, 2007.

Sample

Protein

(%)

Protein digestibility

(%)

Tannin (%)

Phytic acid mg/g

105 days Maturity

4.1a

(±0.01)

39.4a

(±1.1)

4.5b

(±0.07)

0.40b

(±0.02)

90 days Maturity

3.8a

(±0.05)

36.2a

(±1.2)

22.2a

(±0.13)

0.20a

(±0.01)

75 days Maturity

3.7a

(±0.05)

32.2a

(±1.2)

20.2a

(±0.13)

0.15a

(±0.01)

• Mean values of same letters within the column are significantly difference at

(P? 0.05).

About the Author

Murwan K. SabahelKhier and Saifeldin A. Hussain
Department of Biochemistry, School of Biotechnology, Faculty of Science and Technology, Al Neelain University, Sudan.
E-mail: murwansabahelkhier@yahoo.com

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Through the interaction with the resin. If the resin is hydrophobic and you try to separate hydrophobic compounds, the most hydrophobic compound will interact with the resin the strongest so it will be retained on the column the longest; the opposite will be true for a less hydrophobic or polar compound. Also, separation may depend on the size and surface of the resin - small compounds may be retained longer than large ones, because they will partition between the solid phase (the resin) and the mobile phase (the eluent) (size exclusion chromatography). Finally, you may have ionic interactions between the resin and the compounds you want to separate, primarily in ion-exchange chromatography.

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