No abstract

The Hayabusa2 spacecraft collected samples from the surface of the carbonaceous near-Earth asteroid (162173) Ryugu and brought them to Earth. The samples were expected to contain organic molecules, which record processes that occurred in the early Solar System. We analyzed organic molecules extracted from the Ryugu surface samples. We identified a variety of molecules containing the atoms CHNOS, formed by methylation, hydration, hydroxylation, and sulfurization reactions. Amino acids, aliphatic amines, carboxylic acids, polycyclic aromatic hydrocarbons, and nitrogen-heterocyclic compounds were detected, which had properties consistent with an abiotic origin. These compounds likely arose from an aqueous reaction on Ryugu’s parent body and are similar to the organics in Ivuna-type meteorites. These molecules can survive on the surfaces of asteroids and be transported throughout the Solar System.

Chiral amino acid analysis, especially the determination of trace levels of D-enantiomers, is currently gathering attention in a variety of research areas including the food/clinical sciences. These D-amino acids had long been believed to be absent in the higher animals. However, by the advances of analytical technologies, some of the D-enantiomers are found in mammals including humans and increasingly recognized as novel physiologically-active substances and/or biomarkers. For the determination of these D-amino acids and related compounds in real world samples, utilization of sensitive and selective methods is essential and multi-dimensional HPLC is one of the straightforward approaches. In the present review, two/three-dimensional HPLC methods and biological/medical applications focusing on our current studies are summarized.

The concentrations of several D-amino acids have been reported to significantly increase in the plasma of patients with chronic kidney disease (CKD). However, the amounts of these D-amino acids are low (around 1% of the Lform or lower), and their analyses were complicated by various interfering compounds in many clinical samples. A highly selective analytical method is thus required to perform the accurate and precise determination of these D-amino acids in the plasma of CKD patients. In the present study, a selective 3D-HPLC system equipped with reversed-phase, anionexchange, and enantioselective columns has been designed and developed for the determination of the asparagine, serine, alanine and proline enantiomers. For the sensitive analysis, amino acids were precolumn derivatized with 4-fluoro-7-nitro-2,1,3-benzoxadiazole and detected by their fluorescence. The system was validated by using human plasma in addition to the standard amino acids, and results with a sufficient linearity, precision, and accuracy were obtained. The 3D-HPLC system was applied to the plasma of patients with different stages of CKD and all of the target D-amino acids were clearly observed without interferences for all 25 tested patients. Good correlations were shown between the kidney function of the patients and the %D values of the target analytes, especially D-Asn and D-Ser, indicating that the present 3D-HPLC method is useful for the sensitive diagnosis of CKD.

For the enantioselective and simultaneous analysis of lactate and 3-hydroxybutyrate, a validated online two-dimensional high-performance liquid chromatography system using 4-nitro-7-piperazino-2,1,3-benzoxadiazole as a fluorescent derivatization reagent has been developed. For the reversed-phase separation in the first dimension, a Capcell Pak C18 ACR column (1.5 × 250 mm, particle size 3 μm) was used, and the target fractions were isolated by their hydrophobicity. In the second dimension, a polysaccharide-coated enantioselective column, Chiralpak AD-H (2.0 × 250 mm, 5 μm), was used. The system was validated by the calibration curve, intraday precision, interday precision, and accuracy using standards and real human samples, and satisfactory results were obtained. The present method was applied to human plasma and urine, and in the plasma, trace amounts of d-lactate (8.4 μM) and l-3-hydroxybutyrate (1.0 μM), besides high levels of l-lactate (860.9 μM) and d-3-hydroxybutyrate (59.4 μM), were successfully determined. In urine, trace levels of d-lactate (3.7 μM), d-3-hydroxybutyrate (2.3 μM), and l-3-hydroxybutyrate (3.3 μM) in addition to a relatively large amount of l-lactate (15.4 μM) were observed. The present online two-dimensional high-performance liquid chromatography system is useful for the simultaneous determination of all the lactate and 3-hydroxybutyrate enantiomers in human physiological fluids, and further clinical applications are ongoing.