Preparation of Low-Phenylalanine Macro Peptides and Estimation of its Phenylalanine Content by Fluorometric Technique
DOI:
https://doi.org/10.6000/1929-5634.2013.02.03.2Keywords:
Flours, L-phenylalanine, PKU food, dietary supplementationAbstract
The aims of the study were to prepare macro peptides low in phenylalanine (Phe) from non-conventional raw materials, and to demonstrate the feasibility of using the fluorometric technique to measure the diminution of their Phe content. Aqueous solution of flours of legumes, and amaranth panicles were used to elaborate the concentrates by using isoelectric precipitation. These protein concentrates, and a whey solution were incubated with proteolytic enzymes to hydrolyze the peptide link at the aromatic amino acids, and then these macro peptides were filtrated through activated charcoal, in order to reduce its phenylalanine concentration. The Phe concentration, of the each prepared macro peptides, was analyzed by using fluorometric technique, and it was later validated by using HPLC. The crude protein contents in the concentrates have varied from 90% in the protein isolate from lentils, 76% in those from the frijol white, and 44% in those from amaranth panicles. Protein concentrates, and whey were hydrolyzed by using the following enzymes: pepsin from the pig gastric mucosa, protease from Aspergillus oryzae, and protease type XIV from Streptomyces griseus. It was determined that the enzymes with the better hydrolysis capacity, were the proteases from S. griseus and A. oryzae. The macro peptides with non-linked phe were filtered through activated charcoal. Reductions of Phe of up to 99% in the second and third filtrate were observed and this reduction was corroborated by using HPLC technique. It was also established the higher sensitivity of the fluorometric method to detect Phe, than the HPLC technique.
References
FAO Food and Nutrition Series - Collection FAO: Alimentation et nutrition. Amino-acid content of foods and biological data on proteins. FAO Food and Nutrition Series 1981. Available in: http://www.fao.org/docrep/005/ AC854T/AC854T00.htm
Khost NA, Hsieh DST, Shin V, Rha CK. Synthesis of low-Phenylalanine polypeptides. Int J Peptide Protein Res 1982; 20: 267-75. DOI: https://doi.org/10.1111/j.1399-3011.1982.tb03057.x
Arai S, Maeda A, Matsumura M, Hirao N, Watanabe M. Enlarged-scale production of a low-Phenylalanine peptide substance as a foodstuff for patients with Phenylketonuria. Agric Biol Chem 1986; 50(11): 2929-31. http://dx.doi.org/10.1271/bbb1961.50.2929 DOI: https://doi.org/10.1080/00021369.1986.10867855
Lopéz-Bajonero L, Lara-Calderón P, Galvez-Mariscal A, Velázques-Arrellano A, López-Munguia A. Enzymatic production of a low–Phenylalanine product from skim milk powder and caseinate. J Food Sci 1991; 56(4): 938-42. http://dx.doi.org/10.1111/j.1365-2621.1991.tb14610.x DOI: https://doi.org/10.1111/j.1365-2621.1991.tb14610.x
Sigma, 2013. In: http://www.sigmaaldrich.com/us-export.html
Sanvodal M, Ayala S, Ore R. Estimulación de la actividad péptica del jugo gástrico, inducida por látex de Croton palanostigma (sangre de grado). An Fac Med 2008; 69(3): 163-66. DOI: https://doi.org/10.15381/anales.v69i3.1134
Furton JS. The mechanism of catalytic action of pepsin and related acid proteinases. Adv Enzymol 1976; 44: 1-36. DOI: https://doi.org/10.1002/9780470122891.ch1
González EC, Frómeta A, Del Río L, Castells E, Robaina MS, García SM, et al. Cuban neonatal screening of Phenylketonuria using an ultramicro-fluorometric test. Clin Chim Acta 2009; 402: 129-32. http://dx.doi.org/10.1016/j.cca.2008.12.039 DOI: https://doi.org/10.1016/j.cca.2008.12.039
Piecyk M, Śrama A, Bzducha A, Obiedziński M. Application of HPLC AND GC/MS to quantification of Phenylalanine in chosen kinds of food for particular nutritional use. Acta Sci Pol Technol Aliment 2007; 6(2): 5-18.
McCaman MW, Robins E. Fluorometric method for the determination of Phenylalanine in serum. J Clin Med 1962; 59: 885-90.
Wang S, Pizzolato S, Demshar HP. Receiver operating characteristic plots to evaluate guthrie, wallac, and isolab Phenylalanine kit performance for newborn Phenylketonuria screening. Clin Chem 1997; 43(10): 1838-42. DOI: https://doi.org/10.1093/clinchem/43.10.1838
AACC. American Association of Cereal Chemists, Eds. Laboratory Method (10th ed., Methods No. 08-12, 44-15A, 46-13, 30-10, 76-10 and 54-21). St. Paul, Minnesota, USA 2003.
Avanza MV, Puppo MC, Añón MC. Rheological Characterization of amaranth protein gels. Food Hydrocolloid 2005; 19: 889-98. http://dx.doi.org/10.1016/j.foodhyd.2004.12.002 DOI: https://doi.org/10.1016/j.foodhyd.2004.12.002
Bamdad F, Hossein G, Kadivar M. Preparation and characterization of proteinous film from lentil (Lens culinaris) Edible film from lentil (Lens culinaris). Food Res Int 2006; 39: 106-11. http://dx.doi.org/10.1016/j.foodres.2005.06.006 DOI: https://doi.org/10.1016/j.foodres.2005.06.006
Soares R, Biasutti E, Capobiango M, Vieira C, Silva V, Morais H, et al. Preparation of enzymatic skim milk hydrolysates with low phenylalanine content. Acta Farm Bonaerense 2006; 25(3): 325-32.
Abbott D, Andews RS. Introducción a la Cromatografía. 3rd ed. Madrid: Alhambra 1970.
Ault A. Techniques and Experiments for Organic Chemistry. USA: University Science Books 1998.
Gorinstein S, Pawelzik E, Delgado E, Haruenkit R, Weiszy M, Trakhtenberg S. Characterization of pseudo cereal and cereal proteins by protein and amino acid analyses. J Food Sci Agric 2002; 82: 886-89. http://dx.doi.org/10.1002/jsfa.1120 DOI: https://doi.org/10.1002/jsfa.1120
Udenfriend S. Development of a new fluorescent reagent and its Application to the automated assay of amino acids and peptides at the picomole level. J Res Nat Bureau of Standards -A Phys Chem 1972; 76A(6): 638-40. DOI: https://doi.org/10.6028/jres.076A.055
Hui HY, Cross N, Kristisson HG, Lim, MH, Nip WK, Siow LF, Stanfield PS. Biochemistry of seafood processing. In: Simpson BK, editor Food Biochemistry and Food Processing. 2nd ed. Quebec, Canada: Whyley 2012; pp. 109-181. http://dx.doi.org/10.1002/9781118308035.ch19 DOI: https://doi.org/10.1002/9781118308035.ch19
Takase M, Kawase K, Diyosowa I, Ogasa K, Susuki S, Kuroume T. Antigenicity of casein enzymatic hydrolysate. J Dairy Sci 1979; 62: 1570-76. http://dx.doi.org/10.3168/jds.S0022-0302(79)83463-3 DOI: https://doi.org/10.3168/jds.S0022-0302(79)83463-3
Moszczynski, P, Idziac J. Preparation of enzymatic hydrolysates of casein depleted in phenylalanine. Appl Biochem Microbiol 1993; 29(3): 302-306.
Downloads
Published
How to Cite
Issue
Section
License
Policy for Journals/Articles with Open Access
Authors who publish with this journal agree to the following terms:
- Authors retain copyright and grant the journal right of first publication with the work simultaneously licensed under a Creative Commons Attribution License that allows others to share the work with an acknowledgement of the work's authorship and initial publication in this journal.
- Authors are permitted and encouraged to post links to their work online (e.g., in institutional repositories or on their website) prior to and during the submission process, as it can lead to productive exchanges, as well as earlier and greater citation of published work
Policy for Journals / Manuscript with Paid Access
Authors who publish with this journal agree to the following terms:
- Publisher retain copyright .
- Authors are permitted and encouraged to post links to their work online (e.g., in institutional repositories or on their website) prior to and during the submission process, as it can lead to productive exchanges, as well as earlier and greater citation of published work .
