Study of Synthesis and Biological Function on Aminophosphonic Acids

Aminophosphonic Acids 화합물의 생물학적 기능연구

Since ${\beta}-aminoethylphosphonic$ acid was discovered in the living organism, the biosynthesis and biological function of aminophosphonic acids have been extensively studied. The purpose of this project consists in the two parts: 1)the preparation of DL-1-amino-2-phenylethylphosphonic acid (Phenylalanine aminophosphonic acid) and DL-1-amino-3-methylbutyl-phosphonic acid (Isoleucine aminophosphonic acid) by the method of Chamber and Isbell. 2) the study of metabolism and biological functions of those synthetic materials by the animal experiment (white rats) The importance of this project proved to be the first experience fed by animals for the elucidation of biochemical and metabolic functions in the animal body. The following organic synthesis of DL-1-amino-3-methylbutylphosphonic acid and DL-1-amino-2-phenylethylphosphonic acid are studied. 1)Synthesis of DL-1-amino-3-methylbutylphosphonic acid a) Synthesis of Iso-butylbromide b) Synthesis of Ethyl iso-butylmalonate c) Synthesis of Iso-caproic acid d) Synthesis of $Ethyl-{\alpha}-bromo$ iso-caproate e) Synthesis of $Triethyl-{\alpha}-phosphono$ iso-caproate f) Synthesis of DL-1-amino-3-methylbutylphosphonic acid 2)Synthesis of DL-1-amino-2-phenylethylphosphonic acid a) Synthesis of Diethyl phosphite b) Synthesis of Ethylchloro acetate c) Synthesis of Triethyl phospho acetate d) Synthesis of Triethyl benzyl phospho acetate e) Synthesis of DL-1-amino-2-phenylethylphosphonic acid The synthetic compounds; DL-1-amino-3-methylbutylphosphonic acid and DL-1-amino-2-phenyl ethylphosphonic acid which are essential amino acid (isoleucine, phenylalanine)analogue are supplemented to the animal diet at the level of 0.2% and 0.4% for isoleucine analogue and 0.35% and 0.7% for phenylalanine analogue. The plain isoleucine and phenylalanine at the same level in the diet are fercilitated as comparable groups in this study. Two sets of experience including 100 male rats were carried out for seven weeks each total 14 weeks. During this period, urine samples, and each big organs were collected for the analysis of total nitrogen, phosphorus, and glycogen contents in the individual samples by Micro Kjeldahl Fisk & Subbarow and Nelson Somogye, method. 1) The result of the project a) The yield of DL-1-amino-3-methylbutylphosphonic acid and DL-1-amino-2-phenylethylphosphonic acid showed low tendency at the level of 12.5% and 20% Melting point of those two compounds were very high and the ${\alpha}-amino$ group in the synthetic compounds showed positive reaction with ninhydrin in the violet color. b) Ail the experimental groups included in this study revealed statistically no significant difference in the organ weight, total body nitrogen retention and urinary phosphorus excretion This means isoleucine aminophosphonic acid and Phenylalanine aminophosphonic acid were utilized in the body as much as the plain amino acids, isoleucine and phenylalanine did. c) The glycogen contents in the liver of the phenylalaine aminophosphonic acid gruop showed higher statistically significant(p<0.05) in the comparision with the group of the Phenylalanine and the Standard-2. It was noteworthy that the higher glycogen content in the liver might indicate the significance in the incorporation of phenylalanine aminophosphonic acid into the intermediate of tricarboxylic acid cycle as activated state.

Iso-butyl bromide의 합성을 시작으로 D.L-1-amino-3-methylbutylphosphonic acid의 합성을, Diethyl phophite의 합성을 시작으로 D.L-1-amino-2-phenyl ethylphosphonic acid의 합성을 하였다. 합성되어진 필수 아미노산인 Isoleucine analogue phosphonic acid는 각각 0.2%, 0.4%를 식이에 첨가했고 Phenylalanine analogue phosphonic acid는 0.35%와 0.7%를 각각 첨가하였으며 Standard-1과 Standard-2군과 비교군인 Isoleucine 0.2%와 0.4% Phenylalanine 0.35%와 0.7%군으로써 총 100마리의 숫쥐로 각 7주씩 14주의 사육 실험을 하였다. 동물 사육 기간내의 뇨채취와 사육후의 각 장기의 채취로써 총 질소와 P, glycogen측정을 시도하였다. 이러한 일련의 실험결과 (1) D.L-1-amino-3-methylbuthylphosphonic acid나D.L-1-amino-2-phenylethylphosphonic acid의 유기 합성에 있어서는 다 수율이 낮고 상당히 높은 융점을 보였으며 ${\alpha}-NH_2$기가 Ninhydrin 반응에 예민했다. (2) Aminophosphonic acid의 첨가군(D.L-1-amino-3-methylbutyl phosphonic acid, D.L-1-amino-2-Phenylethylphosphonic acid)에서나 Amino acid(Isoleucine, Phenylalanine)첨가군에 있어서 모든 장기의 무게, 총체내질소 보유율, 간장내 총단백질량과 인의양, 및 뇨를 통한 인의 배설량에 각각 큰 차이를 보이지 않았다. 이로 보아서 Phenylalanine Aminophosphonic acid나 Isoleucine aminophosphonic acid가 다 보통 아미노산인 Isoleucine이나 Phenylalanine에 못지않게 체내에서 이용됨이 밝혀졌다. (3) 그러나 Phenylalanine aminophosphonic acid 첨가군이 간의 glycogen 함량에 있어서 Phenylalanine 첨가군과 Standard군과 비교해 보았을때 높았으며 이는 통계적인 유의성을 나타냈다.(<0.05) 이는 Phenylalanine Aminophosphonic Acid가 active state로써 간내에서 쉽게 이용되어서 Tricarboxylic cycle의 intermediate로 incorporate되는 경향으로 생각 할 수 있다.