In situ synthesis during organic analysis of lunar samples

In the development and application of extremely sensitive analytical techniques the need for extreme cleanliness and avoidance of contamination has been generally recognized and complied with in the organic analysis of lunar samples. Much less attention has been paid to the possibility of inadvertent synthesis of certain organic molecules from indigenous smaller organic or even inorganic constituents of the lunar material. At the part-per-billion level of detectability reactions proceeding even at very low yield can lead to detectable products. A particular area of concern should be the amino acid analysis since those substances are known to be formed by condensation of ammonia, cyanide and carbonyl compounds, all potential products of lunar material upon treatment with aqueous media.     

Organic contamination problems in the Viking molecular analysis experiment

The combination of analytical instrumentation selected for the molecular analysis experiment can carry out a survey of the organic compounds present on Mars regardless of their origin. The high sensitivity of this analysis, the limited number of samples which can be analyzed, the close proximity to the landed spacecraft on the surface of Mars which is accessible to the sampling device, the implications of the positive detection of indigenous organic matter in the Martian soil, and our previous experience with meteorites and lunar samples point to the need for a carefully designed program to maintain the inteprity of the analyzed Martian surface samples. A principal problem in interpreting the results of an organic analysis of an extraterrestrial sample is that of distinguishing contaminating material from indigenous material when unknown types and amounts of contaminants make their way into the sample being analyzed. An approach for control of sample integrity in the Viking molecular analysis experiment has been devised which we believe will eliminate such problems. Basically this involves (1) placing an upper limit on the amount of terrestrial contamination that can be tolerated and still allow scientifically meaningful analyses, (2) identifying the potential sources of contamination and analyzing their relative significance, (3) establishing methods to control these sources, and (4) obtaining complete information on the chemical composition of potential contaminants. Our previous experience in the Apollo mission has been of great value in developing the Viking program, perhaps the most important carryover being the recognition of the importance of establishing a comprehensive contamination control program in the early stages of mission planning and hardware design. The upper limit of total allowable organic contamination has been established as 1 μg g^?1. The principal source types, or modes, which contribute to the contamination load have been identified, each requiring a different approach to control. Spacecraft outgassing is controlled by materials selection to minimize outgassing and hermetic sealing whenever possible. Particulate fallout is controlled by selection of materials, particulate seals, cleaning of the spacecraft exterior, and clean room handling. The cleanliness of the direct sample path is controlled by severe materials limitations, ultracleaning, and pressurized sealing of the assembled hardware. Analysis of the relative probabilities of the sources contributing to the allowable contamination and consideration of the practical aspects of achieving a desired level of control for a particular source has resulted in an allocation ‘tree’ whereby fractions of the total allowable contamination are distributed to the various individual sources. These efforts have pointed out the need for more information concerning some of these sources and have actually dictated certain design changes in the spacecraft. Additional information was obtained experimentally on descent engine exhaust characteristics which led to the use of an organically cleaner fuel. In summary, the early recognition in the Viking mission of the importance of organic contamination control has allowed the evolution of a complete contamination control program encompassing spacecraft design, mission operations, flight operations, and the design of the science instrumentation for the molecular analysis experiment.     

Compounds of the organogenic elements in Apollo 11 and 12 lunar samples: A review

Investigations of low molecular weight compounds of the organogenic elements on lunar samples are reviewed. The three general techniques of vacuum pyrolysis, acid hydrolysis, and crushing have been employed by most investigators. Vacuum pyrolysis of lunar fines produce a variety of gaseous species which are either: (1) indigenous, (2) solar wind products and/or (3) chemical reaction products of mineral phases found in the lunar samples. Acid hydrolysis of lunar fines using deuteriumlabeled acids yields evidence for indigenous methane and ethane. Methane and ethane found in the lunar fines are largely derived from the solar wind with only trace amounts indigenous to the samples. Crushing experiments with lunar fines and breccias produce methane, ethane, hydrogen, nitrogen, hydrogen sulfide and the rare gases.     

Aromatic and heteroatom-containing organic compounds in the lunar samples

The data concerning the indigenous organic compounds in the lunar samples has been consistent. The Apollo 11 sample appeared to be moderately contaminated, and most investigators found known terrestrial artifacts in these samples. The Apollo 12 data indicated that perhaps benzene and toluene were indigenous to the Moon at concentrations below 100 parts per billion. Other, more complex organic molecules (in particular of aromatic structure) might also be present, but in concentrations below 1 ppb. In general, the structural types reported have been relatively simple; perhaps indicating that what little organic chemistry occurs on the lunar surface can best be described as reactions between individual atoms of carbon, hydrogen, oxygen and nitrogen.     

Lunar carbon chemistry: Relations to and implications for terrestrial organic geochemistry

The effort expended on the study of lunar carbon chemistry by a sizeable section of the organic geochemical community has resulted in a temporary dislocation in the research output in organic geochemistry. However the long-term beneficial effects of the lunar studies are already obvious in the new and improved methodology, control of contamination, and renewed interest in the characterisation, quantitation and significance of low molecular weight carbon compounds, polymers and inorganic forms of carbon. The role of carbon at the lunar surface is evidently a complex one, the understanding of which should lead to important generalisations for planetary processes.     

Search for amino acids in Apollo returned lunar soil.

The lunar samples from Apollo flights 11 through 17 provided the students of chemical evolution with an opportunity of examining extraterrestrial materials for evidence of early prebiological chemistry in the solar system. Our search was directed to water-extractable compounds with emphasis on amino acids. Gas chromatography, ion-exchange chromatography and gas chromatography combined with mass spectrometry were used for the analysis. It is our conclusion that amino acids are not present in the lunar regolith above the background levels of our investigations.     

Search for amino acids in apollo returned lunar soil

The lunar samples from Apollo flights 11 through 17 provided the students of chemical evolution with an opportunity of examining extraterrestrial materials for evidence of early prebiological chemistry in the solar system. Our search was directed to water-extractable compounds with emphasis on amino acids. Gas chromatography, ion-exchange chromatography and gas chromatography combined with mass spectrometry were used for the analysis. It is our conclusion that amino acids are not present in the lunar regolith above the background levels of our investigations.     

Problems in the search for amino acids in lunar fines

In the search for amino acids in lunar fines, a major problem is the prevention of contamination from terrestrial sources, and the recognition of terrestrial contamination when it has occurred. Synthesis of amino acids conceivably could take place in the lunar module rocket exhaust, a possibility that has not been adequately ruled out. Amino acids could be shed from the astronauts suits, a possibility which has not been studied at all. Amino acids could also be introduced, at many stages of terrestrial manipulation, and during the analytical procedures employed. Hand contamination has qualitative and quantitative features that are characteristic and can be assessed. Precautions for elimination of hand and microbial contamination from glassware, reagents and water are proposed.A second major problem is the efficiency of recovery of amino acids added to lunar material, which is then subject to the complete analytical scheme. This necessitates the availability of lunar material to develop proper procedures.Besides the amino acids present in excess of blank values, it is necessary for the correct interpretation of any positive findings to know whether the amino acids are free or bound and optically inactive or active.The ion exchange chromatography and gas chromatography-mass spectrometry are procedures that complement each other. Both should be applied not only to the same sample but to the same preparations. To pit one method against the other is to risk losing the best analytical data.     

Lunar organic analysis: Implications for chemical evolution

The lunar samples provided some material from the early period of the solar system. These samples could have preserved in them some of the stages in the evolution of carbon compounds. The analysis, so far, has not revealed with certainty a single organic compound of biological significance. However, the finding of methane, carbon monoxide, carbides, etc. could be considered in the context of cosmic evolution.     

Microbial contamination associated with the Apollo 6 spacecraft during final assembly and testing

The National Aeronautics and Space Administration (NASA) requires that microorganisms which could contaminate the surface of the moon as the result of lunar missions be enumerated and identified so that life forms in lunar materials returned to earth may be more easily recognized as being of native or terrestrial origin.Assessment of microbial contamination in the intramural environments used for the assembly and test of the manned lunar spacecraft (Apollo) was made using fallout strips and air samplers. Microbial contamination on the surfaces of Apollo Command and Lunar Modules was determined by use of the swab-rinse method.Preliminary results indicate that the levels of microbial contamination which accumulated on exposed stainless steel surfaces, as well as airborne microbial contamination in the high bay assembly areas, were similar to those encountered in the unmanned spacecraft assembly areas. However, higher levels of microbial contamination were detected on the Apollo spacecraft than on the unmanned lunar spacecraft.     

Distribution and isotopic abundance of biogenic elements in lunar samples

A review of the abundance of six biologically important elements (H, C, N, O. P and S) demonstrates that they are present in trace amounts only in lunar matter analysed to date. To the endogenous lunar content, elements are contributed by solar wind irradiation and meteorite impacts. However, it is not yet possible to determine the relative importance of the three sources. Enrichment of the heavy isotopes, C¹³, O¹⁸, S³⁴ suggest that these elements may be lost from the lunar surface by hydrogen stripping (from solar wind protons) as volatile gases. The general lack of water, suggests that organic synthesis could not easily be accomplished in lunar rocks. High energy irradiation of the lunar surface may result in rapid destruction of organic matter not protected by a silicate matrix. It is apparent from present data available, that the ambient lunar surface could not support metabolism of known microorganisms.Publication No. 997 Institute of Geophysics and Planetary Physics, University of California at Los Angeles, Los Angeles, California 90024.     

EMBO workshop report. An eclipse over the cell nucleus functional organization of the cell nucleus Prague, August 9-12, 1999

The EMBO workshop 'Functional Organization of the Cell Nucleus' held in Prague at the Hotel of the Postgraduate School of Medicine was attended by 110 participants (49 invited speakers and 61 selected participants) from 22 countries. Such a full range of topics devoted to the cell biology of the nucleus has not been discussed previously in such an intimate meeting in Europe. The workshop not only offered an opportunity for junior scientists to benefit from having an international meeting within Europe and a chance to discuss their work with internationally recognized experts, but it also offered a unique opportunity for interactions among the more established investigators. The fruits of a number of presentations are gathered together in a Special Issue of the Journal of Structural Biology which appeared in spring 2000. Last but not least, it is worth mentioning that while stepping through a packed scientific program, the participants did find the time to observe the solar eclipse just before the lunch break of August 11.     

Assessing Genetic Heterogeneity within Bacterial Species Isolated from Gastrointestinal and Environmental Samples: How Many Isolates Does It Take?

Strain typing of bacterial isolates is increasingly used to identify sources of infection or product contamination and to elucidate routes of transmission of pathogens or spoilage organisms. Usually, the number of bacterial isolates belonging to the same species that is analyzed per sample is determined by convention, convenience, laboratory capacity, or financial resources. Statistical considerations and knowledge of the heterogeneity of bacterial populations in various sources can be used to determine the number of isolates per sample that is actually needed to address specific research questions. We present data for intestinal Escherichia coli, Listeria monocytogenes, Klebsiella pneumoniae, and Streptococcus uberis from gastrointestinal, fecal, or soil samples characterized by ribotyping, pulsed-field gel electrophoresis, and PCR-based strain-typing methods. In contrast to previous studies, all calculations were performed with a single computer program, employing software that is freely available and with in-depth explanation of the choice and derivation of prior distributions. Also, some of the model assumptions were relaxed to allow analysis of the special case of two (groups of) strains that are observed with different probabilities. Sample size calculations, with a Bayesian method of inference, show that from 2 to 20 isolates per sample need to be characterized to detect all strains that are present in a sample with 95% certainty. Such high numbers of isolates per sample are rarely typed in real life due to financial or logistic constraints. This implies that investigators are not gaining maximal information on strain heterogeneity and that sources and transmission pathways may go undetected.     

The search for indigenous lunar organic matter

Early lunar conditions are not inconsistent with the production of large quantities of prebiological organic matter now sequestered in the lunar subsurface. The absence of substantial water and organic compounds from the superficial layers of the lunar surface may not be indicative of the composition of deeper regions.     

Review of methods used in lunar organic analysis: Extraction and hydrolysis techniques

Extraction, hydrolysis and crushing procedures have proven useful in analyzing for some of the carbon-containing compounds which are present in Apollo 11 and 12 lunar samples. Three main extraction methods employed with aqueous and nonaqueous solvents were refluxing in open and closed systems, Soxhlet extraction, and sonication. With water and acids, refluxing was the method of choice. Of the various nonaqueous solvent systems used, benzene: methanol mixtures were most often selected, and sonication was favored over Soxhlet extraction. Extraction of lunar samples with water followed by acid hydrolysis of the water extract was found to be superior to direct acid hydrolysis of lunar material in the search for amino acids or their precursors. Direct acid hydrolysis of lunar materials did demonstrate however, that carbides or carbide-like compounds are present on the moon. Hydrolysis with deuterated acids and bases showed that lunar samples contain indigenous methane and ethane and confirmed the presence of carbide-like materials. Crushing experiments also showed that gaseous hydrocarbons can be released from lunar samples.     

Multiple uses of liquepipets in Southern, northern, and western blots

Individually wrapped, sterile disposable transfer pipets can be used in the isolation of ds-DNA and ds-RNA fragments from gels as well as in the screening of multiple samples in Southern, Northern, and Western blots without potential contamination by exogenous nucleases and proteases. The sensitivity and results obtained by this method are comparable to those obtained by conventional methods. All the prehybridization, blocking, hybridization, and detection processes can be performed within the transfer pipet. The isotopically labeled probes used in hybridization can easily be recovered, stored for reuse, or disposed of as waste with no potential contamination of personnel or laboratory equipment. Strip blots are stable in appropriate buffers within the liquepipets which can be shipped easily worldwide for comparative analyses by collaborative investigators. This method is simple, time saving, and inexpensive and is particularly suitable for multiple sample screening. Other potential applications of this procedure are discussed.     

Animal habitats for space experiments.

There has been little opportunity for flight experiments using small animals, due to delay of construction of the International Space Station. Therefore, proposals using small animals have been unfortunately excepted from International Space Life Sciences Experiment application opportunity since 2001. Moreover, NASA has changed their development plan of animal habitats for space experiments according to changes of the U.S. space policy and the outlook is not so bright. However, international researchers have been strongly requesting the opportunity for space experiments using small animals. It will be also important for Japanese researchers to make a request for the opportunity. At the same time, researchers have to make an advance in ground based studies toward space experiments and to respond future application opportunities immediately. In this symposium, we explain the AEM (Animal Enclosure Module), the RAHF (Research Animal Holding Facility), and the AAH (Advanced Animal Habitat). It will be helpful for investigators to have wide knowledge of what space experiment is technically possible. In addition, the sample share program will be introduced into our communities. The program will provide many researchers with the organs and tissues from space-flown animals. We will explain the technical aspect of sample share program.     

Environmental risk assessment of heavy metals pollution in aquatic ecosystem—A case study: Sediment of Kor River,Iran

A comprehensive investigation of fractionation and environmental risk of nine heavy metals is carried out for 12 sediment samples collected from Kor River, Iran. For this purpose, the 5-stage sequential extraction method, along with individual contamination factor, global contamination factor, and Environmental Risk Index (ERI), is used. Total concentrations of Cr, Hg, Ni, and Zn were found to be beyond the threshold effect level. The results of fractionation patterns indicate that As, Cr, Ni, Pb, and Zn are mostly associated with Fe-Mn oxide fraction, organic fraction, and residual fraction, while Cd and Mo are predominantly associated with carbonate fraction. Cu and Hg are mostly associated with organic and exchangeable fractions. The results of ERI revealed High to Dangerous risks in 40% of Kor River stations. The applied approach in this study is beneficial to other environmental studies that require analysis of complex data.     

Determination of deoxynivalenol in organic and conventional food and feed by sol–gel immunoaffinity chromatography and HPLC–UV detection

The paper describes the determination of deoxynivalenol (DON) in 55 wheat food and feed samples, 26 from conventional and 29 from organic production. Immunoaffinity columns prepared by entrapping anti-DON antibodies by the sol–gel method were used for sample clean-up. DON was quantified by high performance liquid chromatography (HPLC) and ultraviolet (UV) detection. In general, the incidence of DON contamination was rather low. In eight samples (14.5%) the DON concentration was above the LOQ (380 ng/g), in six samples (10.9%) DON was detected but could not be quantified (>LOD (200 ng/g), <LOQ). In seven conventional samples (two pasta, two cookie, two snack and one feed sample) but only in one organic sample (a snack) the DON concentration was >LOQ. The data indicate both a higher incidence of DON contamination and higher DON concentrations in food and feed samples from conventional than in those from organic production.     

High mutagenic activity formed in pan-broiled pork

Lean pork was pan-broiled at various temperatures between 100 and 290 degrees C. Cooking was performed in an open frying pan common for domestic use in Sweden. No fat was added. Cooking procedures are clearly defined in order to facilitate inter-laboratory comparisons. The crust was extracted with organic solvents of varying polarity. The mutagenic activity was assayed with Ames' Salmonella mutagenicity test. Large amounts of mutagenic activity were detected in samples pan-broiled at 200-290 degrees C. The mutagenic activity recovered was about 10 times higher than that reported by previous investigators to be found during cooking of meat under similar conditions. This discrepancy could be due to differences in the composition of Swedish pork as compared to the meat samples used by other investigators or to different methodology in cooking and extraction procedures.