The Fate of Amino Acids During Simulated Meteoritic Impact

ASTROBIOLOGY Volume 9, Number 10, 2009 ª Mary Ann Liebert, Inc. DOI: 10.1089=ast.2008.0327

The Fate of Amino Acids During Simulated Meteoritic Impact Maryle`ne Bertrand,1 Sjerry van der Gaast,2 Faith Vilas,3 Friedrich Ho¨rz,4 Gerald Haynes,4 Annie Chabin,1 Andre Brack,1 and Frances Westall1

Abstract

Delivery of prebiotic molecules, such as amino acids and peptides, in meteoritic=micrometeoritic materials to early Earth during the first 500 million years is considered to be one of the main processes by which the building blocks of life arrived on Earth. In this context, we present a study in which the effects of impact shock on amino acids and a peptide in artificial meteorites composed of saponite clay were investigated. The samples were subjected to pressures ranging from 12–28.9 GPa, which simulated impact velocities of 2.4–5.8 km=s for typical silicate-silicate impacts on Earth. Volatilization was determined by weight loss measurement, and the amino acid and peptide response was analyzed by gas chromatography–mass spectrometry. For all compounds, degradation increased with peak pressure. At the highest shock pressures, amino acids with an alkyl side chain were more resistant than those with functional side chains. The peptide cleaved into its two primary amino acids. Some chiral amino acids experienced partial racemization during the course of the experiment. Our data indicate that impact shock may act as a selective filter to the delivery of extraterrestrial amino acids via carbonaceous chondrites. Key Words: Shock—Impact—Meteorite—Organic matter—Racemization—Amino acid. Astrobiology 9, 943–951.

1. Introduction

T

hroughout the history of Earth, impacts have led to the extinction of living organisms, but they have also transported vital molecules essential for life, including organics and water. Extraterrestrial organic matter detected in comets, asteroids, meteorites, and interplanetary dust is considered by many to have played an important role in the chemical processes that led to the emergence of life on Earth (Chyba and Sagan, 1992; Irvine, 1998; Kerridge, 1999; Brack, 2005; Ehrenfreund and Sephton, 2006; Maurette, 2006; Oro et al., 2006). A direct connection between the massive organosynthesis that occurs in interstellar clouds and the presence of prebiotic compounds in primitive planetary bodies was suggested by Epstein et al. (1987) and Kerridge (1986). A large variety of insoluble and soluble organic compounds have been detected in carbonaceous chondrite meteorites, such as Murchison and Murray (Cronin et al., 1993; Pizzarello, 2006; Yabuta et al., 2007). Some of the amino acids in these meteorites and in micrometeorites presented an l-enantiomeric excess, that is, the enantiomeric form of amino acid upon

which all proteins in living organisms are based (Brinton et al., 1998; Pizzarello and Cooper, 2001; Engel et al., 2004; Pizzarello et al., 2004, 2008). The importance of impact events for the delivery of organic matter, especially during the early history of Earth, has been addressed in a few previous studies (Chyba, 1993; Ehrenfreund et al., 2002; Managadze, 2007). Subsequent investigations on the synthesis of organic molecules, especially amino acids, have been made under simulated interstellar environmental conditions (Bernstein et al., 2002; Munoz Caro et al., 2002), and the behavior of amino acids exposed to space conditions is under investigation (Barbier et al., 2000; Boillot et al., 2002; Cottin et al., 2008). However, the ability of biologically important molecules to survive the high temperatures and pressures generated during impact is poorly understood, due mainly to a lack of relevant experimental data. The composition of gas mixtures that may have been released into the atmosphere by devolatilization of an impactor was analyzed by experimental vaporization of silicates (Mukhin et al., 1989). In another experiment, three samples of the Murchison meteorite were shocked up to 36 GPa, which led to devolatilization of

1

Centre de Biophysique Mole´culaire-CNRS affiliated with the University of Orle´ans, Orle´ans, France. Netherlands Institute of Sea Research, Den Burg, the Netherlands. 3 MMT Observatory, University of Arizona, Tucson, Arizona, USA. 4 NASA Johnson Space Center, Houston, Texas, USA. 2

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944 amines and aliphatic compounds, as well as a slight change in the composition of the residue (Tingle et al., 1992). Mimura and Toyama (2005) studied the effects of pressure shocks on polycyclic aromatic hydrocarbons, demonstrating their decomposition to sootlike materials. Among the prebiotic molecules, amino acids are particularly interesting because they are among the most important prebiotic building blocks of primitive life (cf. Brack, 2007). The fate of these molecules during impact shock has been investigated both experimentally and by modeling. Saturated solutions of amino acids were shocked at pressures ranging from 5.1 to 21 GPa and at temperatures from 412 to 870 K by Blank et al. (2001), and alanine survived pressures from 1.23 to 3.67 GPa and temperatures up to 5008C (Chen et al., 2007). Peterson et al. (1997) documented the destruction and racemization of the amino acids proline, aspartic acid, glutamic acid, phenylalanine, norleucine, norvaline, and aminobutyric acid embedded in Murchison and Allende meteorite powder. Finally, computer modeling suggests that some amino acids can survive impacts up to 25 km=s in large, 1–5 km diameter asteroids and comets (Pierazzo and Chyba, 1999). These experiments have provided important insight into the fate of amino acids during impact shock under specific experimental conditions. We have undertaken additional impact shock experiments to test the survival of a different range of amino acids and one dipeptide in a more natural context. For example, in our experiment we embedded the organic components in the clay saponite, a common aqueous alteration product of meteorites ( Jones and Brearley, 2006; Ohnishi and Tomeoka, 2007), because organic molecules have a natural affinity for clay materials. We also used concentrations of the molecules that are closer to those found in carbonaceous chondrites and lower than those used in previous experiments. Nine of the eleven amino acids we used have been identified in the Murchison meteorite (Cronin et al., 1995), and some of them are found in other carbonaceous meteorites (Botta et al., 2002). Given the importance of the lenantiomeric forms of amino acids in proteins and the excesses found in some amino acids in the Murchison and Murray carbonaceous chondrites (Pizzarello and Cooper, 2001; Engel et al., 2004; Pizzarello, 2004) and micrometeorites (Brinton et al., 1998; Pizzarello et al., 2008), we paid special attention to enantio-inversion (partial racemization), that is, the transformation of a given enantiomer into its mirror image during impact shocks. The objective of our investigation was to study experimentally the resistance of amino acids to impact shock as the function of their chemical nature. 2. Materials and Methods All amino acids and dipeptide in the post-impact samples were analyzed by gas chromatography–mass spectrometry after extraction with use of several derivatization methods (Abe et al., 1996; Bertrand et al., 2008). 2.1. Amino acids used in this experiment We used two types of amino acids and a dipeptide for our experiments (Table 1): (i) amino acids with an alkyl side chain [Abu*, Aib*, d-Ala, Gly, Nva*, tLeu*, and d-Val (*the nonproteic amino acids were selected as the (S)-enantiomeric

BERTRAND ET AL. form)]; (ii) amino acids with a functional side chain (d-Asp, d-Cys, d-Ser, and Dpr*); (iii) a dipeptide d-Ala-d-Ala. The d-enantiomers were used to avoid any biological contamination since the l-form is used in almost all biological components. Six of the amino acids are proteic (d-Ala, Gly, d-Val, d-Asp, d-Cys, d-Ser). 2.2. Starting materials Each sample was prepared by mixing 175 mg of the clay saponite (variety of montmorillonite) with a solution of 0.5 mmol of glycine, (Gly), d-alanine, (d-Ala), d-valine (d-Val), d-aspartic acid (d-Asp), d-serine (d-Ser), d-cysteine (d-Cys), aminobutyric acid (Abu), aminoisobutyric acid (Aib), diaminopropionic acid (Dpr), tert-leucine (tLeu), and norvaline (Nva) and d-alanyl-d-alanine (d-Ala2). Pure saponite was used for the control experiments. 2.3. Shock experiments We followed the shock procedure described by Peterson et al. (1997). The amino acids mixed with saponite (sample volumes estimated 0.6 cm3) were placed in a stainless steel well (2.5 cm in diameter, 1.12 cm in depth) that was screwed at its base into an outer ring (3.4 cm in diameter, 2 cm in depth). A plug was screwed on top of the well to compress the sample into a disk of about 0.1 cm in depth and 1.3 cm in diameter. The assembly (well þ ring þ plug) was press-fitted into the recess of a target holder, a large metal cylinder made of stainless steel (10 cm in diameter and 12 cm in length). The target comprised both the cylinder and sample-holder assemblage. A 20 mm powder-propellant gun with its barrel extending into a vacuum chamber (10
2 torr) was used to launch a projectile with a chosen velocity into the target material. The projectile was a cylindrical plug of Lexan
faced with a flat metal plate to produce a planar shock wave in the target. The projectile velocity was measured by using the occultation of infrared laser beams trained onto photodiodes. In turn, this velocity measurement was used to calculate the peak pressures via the known equations of state of both target and projectile materials (aluminum 2024, stainless steel 304, Lexan
). The test specimen embedded into the target equilibrates with the metal jacket and experiences the shock stresses calculated for the container. Shock pressures of 12.0, 15.3, 21.0, or 28.9 GPa were applied. After impact, the target was quenched as rapidly as possible to minimize potential secondary effects of long-term residence at elevated temperatures, and the vacuum chamber was rapidly pressurized for quick recovery of the target cylinder. The cylinder was placed on a massive, precooled (
208C) aluminum block with the impacted side facing the cold surface. This entire procedure took less than 2 min, from the impact to contact with the precooled metal block. The temperatures reached in our experiment were not determined. Peterson et al. (1997) estimated a temperature of 588C at 30 GPa. Following impact, the face of the sample holder was sufficiently deformed so that it could not be unscrewed. Instead, it was machined open while being continuously cooled by a steam of gaseous N2 fed from a nearby dewar. The sample was never touched, even by gloved fingers. This process allowed recovery of the sample without any manual contact and thus eliminated the risk of contamination. The impacted samples

SIMULATED METEORITIC IMPACT

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Table 1. Chemical Formula of the 11 Amino Acids and of the Dipeptide Used Gly Glycine MW: 75 g.mol
1

D-Ser D-Serine MW: 105 g.mol
1

D-Ala D-Alanine MW: 89 g.mol
1

D-Asp D-Aspartic Acid MW: 133 g.mol
1

Aib Amino isobutyric Acid MW: 103 g.mol
1

Dpr Diamino propionic Acid MW: 140 g.mol
1

Abu Amino butyric Acid MW: 103 g.mol
1

D-Cys D-Cysteine MW: 175 g.mol
1

D-Val D-Valine MW: 117 g.mol
1

Nva Norvaline MW: 117 g.mol
1

tLeu tert-Leucine MW: 131 g.mol
1

D,Ala-D,Ala dialanine MW: 160 g.mol
1

were stored in small glass vials at 188C until analysis. Two experiments consisting of four samples each were conducted, but not all experiments were successful. We therefore have results for two samples at 12 and 15 GPa, but only one for 21 and 28.9 GPa. The results shown are the average of the two experiments for 12 and 15 GPa. The control experiment consisted of analyses of amino acids and the dipeptide extracted from a saponite sample that had not been shocked. 2.4. Analyses Particular attention was paid to the amino acid extraction procedure, which was optimized to recover the totality of the post-impact sample without inducing racemization or the formation of derived amino acids during the extraction and analytical processes (Shock and Schulte, 1990). The organic compounds were extracted from the saponite, functionalized to become volatile, and analyzed by gas chromatography coupled to mass spectrometry. 2.4.1. Amino acid extraction. The amino acids were recovered from the clay by three successive extractions with methanol=water solutions (50=50). The clay suspensions were sonicated (15 min) and centrifuged (15 min at 10,000 rpm). The combined supernatants were evaporated by speed vacuum, and the residue dissolved in 1 ml of ultrapure water

under sonication (15 min). The samples were stored in small glass vials at
188C. 2.4.2. Amino acid derivatization. The amino acids were derivatized with the addition of an alkyl chloroformate and an alcohol by the acylation=esterification procedure (Husek, 1998, 2005). This procedure, as described by Abe et al. (1996) and Bertrand et al. (2008), ensures minimal sample handling and no heating or evaporation. Each sample was derivatized by using two different alcohols, and the procedure was repeated for a total of 4 derivatizations. Five successive injections were performed for each derivatized solution (20 injections in total) to verify the reproducibility of the results. All samples were functionalized four times: twice with heptafluorobutanol to form N(O,S)-ethoxycarbonyl heptafluorobutyl ester derivatives, and twice with 2-chloropropanol to form the diastereoisomers of N(O,S)-ethoxycarbonyl 2-chloropropyl ester derivatives of the amino acid enantiomers, which were separated by using an achiral column CPSil 19 CB (Bertrand et al., 2008). For each derivatization, 100 ml of standard solution were placed into a 1 ml microreaction vessel from Supelco, and 5 ml of leucine (from 2.10
4 to 2.10
2 M) were added as the first internal standard then 5 ml of octadecane in hexane (from 10
4 to 10
2 M) as the second internal standard. The amino

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BERTRAND ET AL.

acids were derivatized by adding 34 ml of 2-chloropropanol or 49 ml of heptafluorobutanol and 13 ml of pyridine. Ethyl chloroformate (15 ml) was added to this solution, and the vial was shaken for 10 s. After allowing the CO2 to escape for 5 min, 100 ml of chloroform were added. The vial was shaken vigorously for 2 min, and the derivatives were extracted in the organic layer. Finally, 1 ml of the organic layer was injected into the gas chromatograph. 2.4.3. Gas chromatography. An Agilent 6890 gas chromatograph system, equipped with a CP-Sil 19 CB fused-silica capillary column from Varian (30 m0.25 mm internal diameter, 0.2 mm film thickness), coupled with a 5973 mass spectrometer as detector, was controlled by MSD ChemStation software. Helium was used as the carrier gas (inlet pressure: 170 kPa). Splitless injection mode was used. The injector temperature was set at 2308C and the detector temperature at 2508C. The oven temperature was set at 1208C for 5 min, then programmed to reach 2008C at a rate of 38C=min, and finally held at 2008C for 5 min. 2.4.4. Solvents and reagents. Amino acids were purchased from Sigma or Aldrich. Methanol, pyridine, ethyl chloroformate, trichloromethane were obtained from Fluka. (S)-(þ)-2-chloro-1-propanol 97% was purchased from Aldrich. 3. Results 3.1. Capsule material recovery After experimental shock, the sample material was recovered and placed in individual glass vials. The samples were weighed before extracting the amino acids from the saponite with a methanol=water solution. 3.2. Impact devolatilization The samples were measured for weight loss before and after each experiment. Weight loss increased with impact pressure (and accompanying shock heating) and was interpreted to record devolatilization of the starting material, as: Devolatilization ¼ (massinitial
massimpact )=massinitial with massinitial being the mass of the sample before impact and massimpact the mass after impact. The devolatilization as a function of the shock pressure is shown in Fig. 1. The devolatilization curve corresponds to third-order polynomial kinetics with a good correlation coefficient (R2 ¼ 0.9986) and follows the equation: y ¼ 0:0018x3
0:0061x2 þ 0:1586x Devolatilization was low (