INTRODUCTION:
It is widely accepted that the origin of life processes began with the formation of important biomonomers from simple molecules present in the prebiotic environment. That the earth had a reducing atmosphere in its early stages was proposed by Oparin1 and his arguments being extended by Bernal2 and Urey3. It has been accepted that the reducing primordial earth atmosphere consisted of hydrides of carbon, nitrogen and oxygen. The earth wrapped in a heavy blanket of all these hydrides was continuously acted upon by various available energy sources4-5. Metal carbides and nitriles might have afforded hydrocarbons and ammonia respectively on reacting with water. Thus the primordial atmosphere of the earth mainly comprised of CH4, CH3, H2 and H2O as well as H2S.
Initially the earth’s temperature was very high which might have been due to frequent volcanic eruptions as well as heavy flux of solar radiation. Gradually, as the earth cooled, the organic compounds which were formed in relatively higher amount in the atmosphere were carried to the earth’s crust by condensation of water vapors and accumulated into pools and finally in oceans. Due to repeated evaporation and condensation of rain droplets the temperature of the earth crust cooled to about 10000C or lower and the resulting highly reactive free radicals .CH, .CH2, .CH3, .NH2, .H, .OH etc formed from CH4, NH3, H2O vapors on intense photolysis as well as thermal fragmentation could have recombined to form a variety of saturated and unsaturated hydrocarbons as follows-
.CH + .CH à CH CH
2CH4 à HC CH +3CH2
Metal carbides might have reacted with steam forming hydrocarbons.
CaC2 + H2O à HC CH + CaO
As the primitive atmosphere was highly reducing the nitrogen would have been transformed into ammonia as a stable compound.
N2+ 3H2 à 2NH3 (Ammonia)
Thus the making and breaking processes resulted in the formation of a number of simple molecules. As soon as liquid water appeared on the surface of the primitive earth, clay minerals probably accumulated on the surface and suspended into deep sea. It has been emphasized by earlier workers6-7that, clays, minerals and silicates might have played a vital role in chemical evolution and it has been suggested that clays near the hydrosphere-lithosphere interface might have adsorbed micro biomonomers on and between their silicate layers, thereby, protecting them from destructive radiations from the sun.
Synthesis of biomolecules from reaction system comprising of acetylene-ammonia-water vapour under prebiotic earth conditions may throw some light on the process of chemical evolution. It has been considered of interest therefore, to study the effect of heat on reaction system of acetylene, ammonia and water vapour to investigate the formation of amino acids in the presence of silica and alumina widely distributed near the sea shores under wetting-drying cycles of primitive earth.
EXPERIMENTAL:
All the investigations were carried out in aqueous medium. The vessels, experimental solutions and pressure for 30 minutes prior to their use in experiments. Sterilized double distilled water was used as the solvent in every experiment wherever necessary the vapors of the double distilled water was allowed to pass through the reaction vessels.
Ammonia and acetylene gases were freshly prepared by the usual methods described in literature. Every care was taken to ensure the purity of the gaseous samples employed. All these gases, when used in the experiments were washed by passing through traps before allowing them to pass through the solutions placed on hot plates.
Ammonia gas was prepared by slowly heating high grade extra pure ammonia liquor of Qualigens (A.R. Grade) make. Acetylene gas was prepared by adding water drop by drop to calcium carbide. The evolved gas was passed through a number of gas washing traps containing separately a solution of copper acetate, dilute sulphuric acid and finally it was washed through water. The gas thus purified was allowed to bubble through the solutions contained in several glass reaction vessels.
Experimental solution (5 ml each) with pH 9.0 ± 0.5 maintained at a pressure of 20 ± 5 mm/Hg were heated in borosil glass reaction vessels- Kjeldhal flasks (100 ml) fitted with air condensers on hot plates at a temperature of 90 ± 50C in the presence and absence of silica and alumina.
Samples of reaction concentrates were analyzed for the possible formation of amino acids using chromatographic techniques8-9on Whatman No- 01 paper both by uni- and two dimensional chromatography using butanol- acetic acid- water (4:1:1 v/v, 4:1:5 v/v upper layer), butanol- acetic acid- pyridine- water (15:3:10:12 v/v) and phenol- water (80:20 v/v). Amino acid spots were visualized with ninhydrin, identified with isatin and also by comparison of their Rf values with authentic amino acids as well as of their DNP derivatives. For the separation of DNP amino acids the solvent system used was n- butanol saturated with water. Colorimetric estimation of amino acids was carried out by comparison of color intensity of the unknown compound with that of a standard solution employing photochemical colorimeter MK III.
Ultraviolet absorption spectra of the various reaction mixtures or elutes of some products were determined in aqueous solution using Jasco V- series spectrophotometer. IR spectra of reaction concentrates were recorded in Perkin Elmer 881 (4000-6000 cm-1) spectrophotometer.
The reaction products were further identified by High Performance Liquid Chromatography which were ascertained by Shimadzu SPD- 10 A UV visible detector with C18 column using triple distilled water: methanol (80:20 v/v) and 0.1% H3PO4: acetonitrile (40:60 v/v) as mobile phase, flow rate 1.5 ml/min at pH 7.0, temperature 250C and UV detector at 197-210 nm for the detection of amino acids. Results were compared with retention times of the standard amino acids run in the same column under similar conditions
RESULTS AND DISCUSSION:
The effect of heat on mixed gases comprising of acetylene and ammonia in the presence of water vapour has been carried out with a view to synthesize biologically significant molecules especially amino acids under simulated primitive earth sea-beach conditions. Therefore attempts have been made for the possible synthesis of amino acids in the presence and absence of silica and alumina separately under prebiotic wetting and drying condition.
The reaction system of acetylene, ammonia and water heated for a period of 10 hrs and on subsequent chromatography showed no ninhydrin positive products, whereas on prolonging the duration of heating for 25 hrs showed the formation of five ninhydrin positive products in trace amount. [Fig. 1 ] Extending the period of the effect of heat up to 50 hrs showed the formation of eight products on the chromatogram. Out of these, lysine, aspartic acid and serine were formed in trace amount while valine and leucine in relatively moderate amount [Fig.1]. When the effect of heat on reaction system of acetylene, ammonia and water vapor was carried out for 75 hrs under similar conditions in all ten amino acids appeared on the chromatogram. However, the yield of products valine and leucine seem to increase considerably. (Fig. 1 )
Fig.1 Chromatogram showing the formation of Amino Acids from reaction system of Acetylene, Ammonia and water vapor heated up to 100 hours.
On prolonging the duration of heating for 100 hrs showed in all nine ninhdrin positive products on the chromatogram.Glumatic acid ,α alanine, amino butyric acid and valine were formed in appreciable amount while lycine,aspartic acid and serine were formed in moderate amount [Fig2]
Fig.2 Chromatogram showing the formation of Amino Acid in presence of Silica and Alumina heated up to 100 hrs.
Fig. 03 UV absorption spectra of reaction concentrate of Acetylene, Ammonia and Water vapor heated up to 100 hrs.