Organic syntheses from CH4−N2 atmospheres: Implications for Titan

Numerous experiments have already been performed, simulating the evolution of gaseous mixtures containing CH₄ when submitted to energy flux. From their results, it appears that a variety of organic compounds, including unsaturated hydrocarbons and nitriles such as HCN, can be synthesized into noticeable amounts from CH₄–N₂ mixtures. In particular, systematic studies of the influence of the composition of the mixture on the nature and amount of synthesized compounds show that organic volatile nitriles, and particularly cyanoacetylene and cyanogen, are formed only in media rich in nitrogen. Those nitriles have been identified very recently in the atmosphere of Titan, and thus, data from such laboratory experiments may provide important indirect information on the organic chemistry occuring at the periphery of this satellite of Saturn. However, during these experiments, there is a continuous formation and accumulation of molecular hydrogen, which does not occur in the atmosphere of Titan, because of H₂ escape. In order to reassess the data already available from this type of laboratory studies, experiments on CH₄–N₂ atmospheres, with and without H2 escape, have been recently performed. The influence of this parameter on the chemical evolution of the atmosphere and on the nature and relative quantities of organic compounds has been studied.After reviewing these experiments, implications of the obtained results on the organic chemistry at the periphery of Titan are discussed.Paper presented at the 6th College Park Colloquium, October 1981.     

Evolution of asymbiotic nitrogen fixation

Recent observations on the nature of the enzyme complex, nitrogenase, prepared from a variety of nitrogen-fixing micro-organisms, on its substrate specificity, energy requirements, source of reducing power and sensitivity to O₂ now permit speculation on the evolution of biological nitrogen fixation in asymbiotic micro-organisms.Ability to fix N₂ is restricted to procaryotic organisms and is particularly widespread among those having characteristics (e.g. hydrogenase, ferredoxin) regarded as primitive. If the primitive environment was devoid of O₂, the earliest N₂-fixing prokaryote would have been a strict anaerobe, not unlike Clostridium pasteurianum. Yet N₂-fixation seems unnecessary in a primitive ammonia-containing environment, and ammonia represses this function in contemporary species. This apparent paradox, the development of the ability to fix N₂ in circumstances in which it was apparently unnecessary suggests that a substance other than N₂ might have been primary substrate of the primeval enzyme.Substances such as acetylene, cyanide, cyanogen, nitriles or isonitriles are all substrates for nitrogenase and are all probable components of the primitive terrestrial environment. Biologically useful functions which a nitrogenase-like reductase system might have served involving substrates other than N₂ include: (a) a detoxification reaction to nullify the effects of cyanide or cyanogen; (b) a means of generating ATP anaerobically; (c) a hydrogen “escape valve”.Functions (b) and (c) are improbable because they would be physiologically uneconomic; function (a) is plausible.With the emergence of an oxidizing atmosphere, facultative and aerobic N₂-fixing micro-organisms could only retain the nitrogenase system if the O₂-sensitive component was protected from inactivation. In the Azotobacteraceae this is achieved by “conformational protection” together with a high respiration rate; in blue-green algae, a structural compartmentation occurs in the more highly evolved species.     

Far UV irradiation of model prebiotic atmospheres

UV light has been the most important energy source on the primitive Earth. However, very few experiments have been performed to test directly the possible role of this energy source on the chemical evolution of the primitive atmosphere, mainly on account of experimental difficulties. Experiments are generally performed with other excitations, mainly electric discharge, and it is frequently assumed that UV irradiation would give similar results.As theoretical considerations make this assumption questionable, direct experimental controls have been undertaken: Model primitive atmospheres have been submitted to 147 nm UV light and the gaseous phase has been analysed. Preliminary qualitative results concerning CH₄–NH₃ atmospheres are reported.Irradiation of pure CH₄ gives rise to the synthesis of a large number of hydrocarbons, mainly saturated hydrocarbons but including also unsaturated ones as, C₂H₂, C₂H₄, C₃H₆, C₃H₄. These insaturated hydrocarbons are synthetized at a very low rate when ammonia is present in the medium.Irradiations of CH₄–NH₃ mixtures give rise, in addition to hydrocarbons, to important amounts of HCN (about 0.1%) and to lesser amounts of CH₃CN and C₂H₅CN. No unsaturated nitriles such as acrylonitrile and cyanoacetylene have been detected. Search for amines is in progress.These results evidence that UV irradiation may contribute largely to synthesis of HCN in CH₄–NH₃ atmospheres and, consequently to the synthesis of many biochemical compounds that can be derivated from HCN. However, synthesis of other compounds, such as pyrimidines, which can derivate from other nitriles, such as cyanoacetylene, cannot be initiated only by UV light, contrary to electric discharges. In addition, if electric discharges are very efficient for synthesis of nitriles in CH₄–N₂ atmospheres, there is not yet evidence that UV light is able to do so.Presented at the 2nd ISSOL Meeting and the 5th ICOL in Kyoto, 5–10 April, 1977.     

The atmosphere of Titan

Summary The discovery that Titan had an atmosphere was made by the identification of methane in the satellite's spectrum in 1944. But the abundance of this gas and the identification of other major constituents required the 1980 encounter by the Voyager 1 spacecraft. in the intervening years, traces of C₂H₂, C₂H₄, C₂H₆ and CH₃D had been posited to interpret emission bands in Titan's IR spectrum. The Voyager infrared Spectrometer confirmed that these gases were present and added seven more. The atmosphere is now known to be composed primarily of molecular nitrogen. But the derived mean molecular weight suggests the presence of a significant amount of some heavier gas, most probably argon. It is shown that this argon must be primordial, and that one can understand the evolution of Titan's atmosphere in terms of degassing of a mixed hydrate dominated by CH₄, N₂ and³⁶Ar. This model satisfactorily explains the absence of neon and makes no special requirements on the satellite's surface temperature. The organic chemistry taking place on Titan today invites comparision with chemical evolution on the primitive Earth prior to the origin of life.Adapted in commemoration of the many contributions of Harold Urey to the study of planetary atmospheres from an article in press in J Planet Sci (1982)     

Study on the chemical evolution of low molecular weight compounds in a highly oxidized atmosphere using electric discharges

The molecular basis for the chemical evolution of low molecular compounds was studied using electric discharges on a higly oxidized atmosphere comprised of CO₂, N₂ and H₂O. In the gas phase, O₂ and CO were formed by the decomposition of CO₂ and their yields were enhanced by the addition of N₂ to the gas mixture. It was demonstrated that H₂O suppressed the reduction of CO₂ while H₂O also had a role in producing organic compounds such as formic acid and formaldehyde. Infrared analysis of the water soluble products and the inner surface of the reaction vessel indicated the production of compounds more complex than formic acid and formaldehyde. These compounds contained the chemical bonds which were identified to be OH, CO, CN and/or CC.     

The role of sulphur in chemical evolution

Summary Sulphur may have played an important role, mainly as an energy converter, during the initial steps of Chemical Evolution.In atmospheric processes, sulphur, in the form of H₂S might have been a primary energy acceptor and a source of hot hydrogen atoms. The presence of H₂S in the primeval earth atmosphere with a molar ratio of about 10^–2 could have allowed the formation of several volatile S-containing compounds without inhibiting the synthesis of the reactive products which are formed in the absence of H₂S. An evaluation of the quantity of H₂S which could have been included in the primeval atmosphere suggests that such a molar ratio may have been reached.In the primitive soup, the thiols and sulphides formed in the gaseous phase may have evolved, giving rise to various prebiotic syntheses. Studies on the addition reaction of alkanethiols on malonic nitriles in aqueous solutions show two different condensation processes: the formation of thioethers and the formation of iminothioesters. Taking into account the values of the specific rate constants for the two reactions, it is shown that these reactions may have taken place in the primitive earth conditions. These two compounds may have played an important role in the prebiochemical evolution. In particular, iminothioesters can be considered as the immediate precursors of thioesters.     

The role of the atmosphere in nitrogen cycling

In this work an analysis of the sources, atmospheric concentration, chemical reactions and sinks of the principal atmospheric nitrogen compounds is made. Atmospheric emissions of N₂O and NH₃ are almost entirely due to biological activity on the continents and in the oceans. The combustion of fossil fuels and biomass is the principal source of NO_x. The only relevant chemical transformations in the troposphere are the oxidation of NO_x to NO₃ ^? and the formation of ammonium salts. Only 10% of the NH₃ emitted is oxidized. Washout of NH₄ ⁺ and NO₃ ^? by rainfall is the principal mechanism for removing nitrogen compounds from the atmosphere. Part of the N₂O enters the stratosphere and part must be removed in the biosphere by processes not yet established. NO_x produced in the atmosphere by the burning of fossil fuels and biomass and by lightning represents between 30 and 40% of the total nitrogen fixed. A complete nitrogen balance for the troposphere is presented. Since the photochemical oxidation of NO_x is rapid and atmospheric transport is relatively slow with respect to the cycling of water in the troposphere, nitrogen compounds return to the earth's surface close to where they were emitted. Fixed-nitrogen inputs to the continents and oceans due to biological and industrial fixation are slightly greater than those due to rain water. However, since rain falls everywhere, input from this source is only important on soils not subject to intensive agriculture.     

Natural hydrazine-containing compounds: Biosynthesis,isolation, biological activities and synthesis

Hydrazine, hydrazone and hydrazide derivatives are nitrogen–nitrogen bond containing compounds. Such molecules are relatively scarce in nature and have been isolated from plants, marine organisms and microorganisms. These compounds exhibit remarkable structural diversity and relevant biological activities. The enzymes involved in the formation of the N–N bond are still unknown, but many lines of evidence support the involvement of N-nitrosation and N-hydroxylation activating steps. Beside the challenging N–N bond, N-acylases catalyzing the C–N bond formation contribute to the chemical diversity of N–N-containing natural products (N2NP). This review examines the state of knowledge regarding the biosynthesis of N2NP, for which only two biosynthetic gene clusters have been investigated. Biological properties and chemical synthesis of hydrazines, hydrazones and hydrazides are also reported.     

The role of cometary particle Coalescence in chemical evolution

Important prebiotic organic compounds might have been transported to Earth in dust or produced in vapor clouds resulting from atmospheric explosions or impacts of comets. These compounds coalesced in the upper atmosphere with particles ejected from craters formed by impacts of large objects. Coalescence during exposure to UV radiation concentrated organic monomers and enhanced formation of oligomers. Continuing coalescence added material to the growing particles and shielded prebiotic compounds from prolonged UV radiation. These particles settled into the lower atmosphere where they were scavenged by rain. Aqueous chemistry and evaporation of raindrops containing nomomers in high temperature regions near the Earth's surface also promoted continued formation of oligomers. Finally, these oligomers were deposited in the oceans where continued prebiotic evolution led to the most primitive cell. Results of our studies suggest that prebiotic chemical evolution may be an inevitable consequence of impacting comets during the late accretion of planets anywhere in the universe if oceans remained on those planetary surfaces.     

Nitrogen fixation as evidence for the reducing nature of the early biosphere

Probably the first nitrogen fixers were anaerobic, non-photosynthetic, bacteria, i.e. fermenters. During the evolution of N₂ fixation they still needed nitrogen on the oxidation level of ammonia. Because of the complexities in structure and function of nitrogenase this evolution must have required a long time. The photosynthetic and later the respiring bacteria inherited the capacity for N₂ fixation from the fermenters, but the process did not change a great deal when it was taken over.Because of the long need for NH₃, which is unstable in a redoxneutral atmosphere, a long-persisting reducing atmosphere was needed. The transition to a redoxneutral atmosphere, dominated by CO₂, H₂O and N₂, cannot have been rapid, and the NH₃ in it was recycled. Probably the atmosphere contained for a long time, as was suggested by Urey but is often denied now, a great deal of methane as a reductant. The recycling occurred with participation of intermediates like cyanide, through energy input as UV radiation or as electric discharges. A stationary state was set up.The hypothesis is recalled that coloured, photosynthetic, NH₃ bacteria, analogous to coloured sulphur bacteria, may have existed, or may still exist, in reducing conditions. A few remarks are made about the origin of nitrification in the later, oxidizing atmosphere.     

The prebiological paleoatmosphere: Stability and composition

In the past, it was generally assumed that the early atmosphere of the Earth contained appreciable quantities of methane (CH₄) and ammonia (NH₃). This was the type of atmosphere believed to be the most suitable environment for chemical evolution, the nonbiological formation of complex organic molecules, the precursors of living systems. Photochemical considerations suggest that a CH₄–NH₃ dominated early atmosphere was probably very short-lived, if it ever existed at all. Instead, an early atmosphere of carbon dioxide (CO₂) and nitrogen (N₂) is favored by photochemical as well as geological and geochemical considerations. Photochemical calculations also indicate that the total oxygen column density of the prebiological paleoatmosphere did not exceed 10^–7 of the present atmospheric level.Paper presented at the 6th College Park Colloquium, October 1981     

Prebiotic Synthesis of Methionine and Other Sulfur-Containing Organic Compounds on the Primitive Earth: A Contemporary Reassessment Based on an Unpublished 1958 Stanley Miller Experiment

Original extracts from an unpublished 1958 experiment conducted by the late Stanley L. Miller were recently found and analyzed using modern state-of-the-art analytical methods. The extracts were produced by the action of an electric discharge on a mixture of methane (CH₄), hydrogen sulfide (H₂S), ammonia (NH₃), and carbon dioxide (CO₂). Racemic methionine was formed in significant yields, together with other sulfur-bearing organic compounds. The formation of methionine and other compounds from a model prebiotic atmosphere that contained H₂S suggests that this type of synthesis is robust under reducing conditions, which may have existed either in the global primitive atmosphere or in localized volcanic environments on the early Earth. The presence of a wide array of sulfur-containing organic compounds produced by the decomposition of methionine and cysteine indicates that in addition to abiotic synthetic processes, degradation of organic compounds on the primordial Earth could have been important in diversifying the inventory of molecules of biochemical significance not readily formed from other abiotic reactions, or derived from extraterrestrial delivery.     

Nitrogen Fixation By Corona Discharge On The Early Precambrian Earth

We report the first experimental study of nitrogen fixation by corona discharge on the anoxic primitive Earth. The energy yields of nitric oxide (NO) and nitrous oxide (N₂O) were experimentally determined over a wide range of CO₂-N₂ mixtures simulating the evolution of the Earth's atmosphere during the Hadean and Archean eras (from 4.5 ba to 2.5 ba). NO, the principal form of fixed nitrogen in lightning and coronal discharge in early Earth, is produced ten times less efficiently in the latter type of electrical discharge with an estimated maximum annual production rate of the order of 10¹⁰ g yr⁻¹. For N₂O the maximum production rate was estimated to be ∼10⁹ g yr⁻¹. These low rates of syntheses indicate that corona discharges as point discharges on the clouds and ground did not play a significant role in the overall pool of reactive nitrogen needed for the emergence and sustainability of life.     

Increase of Nitrogenase Activity in the Blue-Green Alga Nostoc muscorum (Cyanobacterium)

Preincubation of the blue-green alga (cyanobacterium) Nostoc muscorum under hydrogen or argon (nongrowing conditions, neither CO₂ nor N₂ or bound nitrogen present) in the light resulted in a two- to fourfold increase of light-induced hydrogen evolution and a 30% increase of acetylene reduction. Preincubation under the same gases in the dark led to a decrease of both activities. Cultivation of algae under a hydrogen-containing atmosphere (N₂, H₂, CO₂) increased neither hydrogen nor ethylene evolution by the cells. Formation of both ethylene and hydrogen is due to nitrogenase activity, which apparently was induced by the absence of N₂ or bound nitrogen and not by the presence of hydrogen. Inhibitors of protein biosynthesis prevented the increase of nitrogenase activity. Hydrogen uptake by the cells was almost unaffected under all of these conditions. With either ammonia or chloramphenicol present, nitrogenase activity decreased under growing conditions (i.e., an atmosphere of N₂ and CO₂). The kinetics of decrease were the same with ammonia or chloramphenicol, which was interpreted as being due to rapid protein breakdown with a half-life of approximately 4 h. The decay of nitrogenase activity caused by chloramphenicol could be counteracted by nitrogenase-inducing conditions, i.e., by the absence of N₂ or bound nitrogen. A cell-free system from preconditioned algae with an adenosine 5′-triphosphate-generating system exhibited the same increase or decrease of nitrogenase activity as the intact cell filaments, indicating that this effect resided in the nitrogenase complex only. We tentatively assume that not the whole nitrogenase complex, but merely a subunit or a special protein with regulatory function, is susceptible to fast turnover.     

A Reassessment of Prebiotic Organic Synthesis in Neutral Planetary Atmospheres

The action of an electric discharge on reduced gas mixtures such as H₂O, CH₄ and NH₃ (or N₂) results in the production of several biologically important organic compounds including amino acids. However, it is now generally held that the early Earth’s atmosphere was likely not reducing, but was dominated by N₂ and CO₂. The synthesis of organic compounds by the action of electric discharges on neutral gas mixtures has been shown to be much less efficient. We show here that contrary to previous reports, significant amounts of amino acids are produced from neutral gas mixtures. The low yields previously reported appear to be the outcome of oxidation of the organic compounds during hydrolytic workup by nitrite and nitrate produced in the reactions. The yield of amino acids is greatly increased when oxidation inhibitors, such as ferrous iron, are added prior to hydrolysis. Organic synthesis from neutral atmospheres may have depended on the oceanic availability of oxidation inhibitors as well as on the nature of the primitive atmosphere itself. The results reported here suggest that endogenous synthesis from neutral atmospheres may be more important than previously thought. Stanley L. Miller died May 20, 2007.     

Formation of prebiochemical compounds in models of the primitive earth's atmosphere. II: CH4 - H2S atmospheres.

In order to understand the role of sulfur in the primitive atmosphere, we have studied the action of a silent discharge on mixtures of CH4 and H2S at low pressure. The nature of the products formed in the gaseous phase, and the influence of several parameters, especially the H2S percentage, on the yield of the products are reported. The analysis of the products is carried out by gas liquid chromatography and infrared spectrometry. The formation of sulfur-containing compounds, such as thiols and sulfides, is reported. CS2 is formed in high yield (a few percent) in mixtures containing 40-50% of H2S, while the maximum concentration of thiols (i.e., CH3SH and C2H5SH) is reached with lower percentages of H2S. The formation of hydrocarbons decreases rapidly with increasing proportions of H2S. These results show the important inhibitor effect of H2S on the formation of hydrocarbons and the possibility of occurrence of many sulfur compounds in prebiological evolution.     

The Use of Ascorbate as an Oxidation Inhibitor in Prebiotic Amino Acid Synthesis: A Cautionary Note

It is generally thought that the terrestrial atmosphere at the time of the origin of life was CO₂-rich and that organic compounds such as amino acids would not have been efficiently formed abiotically under such conditions. It has been pointed out, however, that the previously reported low yields of amino acids may have been partially due to oxidation by nitrite/nitrate during acid hydrolysis. Specifically, the yield of amino acids was found to have increased significantly (by a factor of several hundred) after acid hydrolysis with ascorbic acid as an oxidation inhibitor. However, it has not been shown that CO₂ was the carbon source for the formation of the amino acids detected after acid hydrolysis with ascorbic acid. We therefore reinvestigated the prebiotic synthesis of amino acids in a CO₂-rich atmosphere using an isotope labeling experiment. Herein, we report that ascorbic acid does not behave as an appropriate oxidation inhibitor, because it contributes amino acid contaminants as a consequence of its reactions with the nitrogen containing species and formic acid produced during the spark discharge experiment. Thus, amino acids are not efficiently formed from a CO₂-rich atmosphere under the conditions studied.     

The geobiological nitrogen cycle: From microbes to the mantle

Nitrogen forms an integral part of the main building blocks of life, including DNA, RNA, and proteins. N₂ is the dominant gas in Earth's atmosphere, and nitrogen is stored in all of Earth's geological reservoirs, including the crust, the mantle, and the core. As such, nitrogen geochemistry is fundamental to the evolution of planet Earth and the life it supports. Despite the importance of nitrogen in the Earth system, large gaps remain in our knowledge of how the surface and deep nitrogen cycles have evolved over geologic time. Here, we discuss the current understanding (or lack thereof) for how the unique interaction of biological innovation, geodynamics, and mantle petrology has acted to regulate Earth's nitrogen cycle over geologic timescales. In particular, we explore how temporal variations in the external (biosphere and atmosphere) and internal (crust and mantle) nitrogen cycles could have regulated atmospheric pN₂. We consider three potential scenarios for the evolution of the geobiological nitrogen cycle over Earth's history: two in which atmospheric pN₂ has changed unidirectionally (increased or decreased) over geologic time and one in which pN₂ could have taken a dramatic deflection following the Great Oxidation Event. It is impossible to discriminate between these scenarios with the currently available models and datasets. However, we are optimistic that this problem can be solved, following a sustained, open‐minded, and multidisciplinary effort between surface and deep Earth communities.     

Ultrasound-Induced Formation of S-Nitrosoglutathione and S-Nitrosocysteine in Aerobic Aqueous Solutions of Glutathione and Cysteine

S-Nitrosocompounds are formed when aqueous solutions of cysteine or glutathione are exposed to ultrasound (880 kHz) in air. The yield of the S-nitrosocompounds was as high as 10% for glutathione and 4% for cysteine of the initial thiol concentrations (from 0.1 to 10 mM) in the aqueous solutions. In addition to the formation of S-nitrosocompounds, thiol oxidation to disulfide forms was observed. After the oxidation of over 70% of the sulfhydryl groups, formation of peroxide compounds as well as cysteic acid derivatives was recorded. The formation of the peroxide compounds and peroxide radicals in the ultrasound field reduced the yield of S-nitrosocompounds. S-Nitrosocompounds were not formed when exposing low-molecular-weight thiols to ultrasound in atmospheres of N₂ or CO. In neutral solutions, ultrasound-exposed cysteine or glutathione released NO due to spontaneous degradation of the S-nitrosocompounds. N₂O₃, produced due to the spontaneous degradation of the S-nitrosocompounds in air, nitrosylated sulfhydryl groups of glutathione manifested in the appearance of new absorption bands at 330 and 540 nm. The nitrogen compounds formed in an ultrasound field modified the sulfhydryl groups of apohemoglobin and serum albumin. The main target for ultrasound-generated oxygen free radicals were cystine residues oxidized to cysteic acid residues.     

The evolution of nitrogen cycling

Based upon arguments concerning properties of the environment and the energetics of nitrogen transformation reactions, new hypotheses regarding their evolution are presented. These hypotheses are supported by new calculations and observations germane to understanding the evolution of the nitrogen cycle. From calculations of shock production by meteor impact, we suggest that impact produced fixed nitrogen could have resulted in the entire reservoir of Earth's N₂ being converted into fixed nitrogen at the end of accretion. We have significantly improved upon previous calculations of the abiotic fixation rate on the early earth and find a rate of fixation by lightning of 1–3 × 10¹⁶ Molecules NO/J, which is 2 to 3 times greater than previous estimates. This strengthens the suggestion, corroborated by the predominance of a single nitrogenase enzyme, that biological nitrogen fixation may have been a late evolutionary development, after the development of an aerobic atmosphere. In addition, we show for the first time that HNO, predicted to be the main product of atmospheric photochemical reactions involving NO on the primitive Earth by photochemical models, would eventually become NO₂ ^– and NO₃ ^– after reaching the Earth's surface. Based upon microbe-environment interactions on an ecological as well as a biochemical scale we suggest that denitrification arose prior to aerobic respiration and that nitrification arose after the advent of an aerobic atmosphere. We hypothesize the following evolutionary sequence for the biological transformation of nitrogen compounds: Ammonification Denitrification Nitrification Nitrogen fixation.