Revision of the Absolute Configurations of Chelocardin and Amidochelocardin.

Even with the aid of the available methods, the configurational assignment of natural products can be a challenging task that is prone to errors, and it sometimes needs to be corrected after total synthesis or single-crystal X-ray diffraction (XRD) analysis. Herein, the absolute configuration of amidochelocardin is revised using a combination of XRD, NMR spectroscopy, experimental ECD spectra, and time-dependent density-functional theory (TDDFT)-ECD calculations. As amidochelocardin was obtained via biosynthetic engineering of chelocardin, we propose the same absolute configuration for chelocardin based on the similar biosynthetic origins of the two compounds and result of TDDFT-ECD calculations. The evaluation of spectral data of two closely related analogues, 6-desmethyl-chelocardin and its semisynthetic derivative 1, also supports this conclusion.

Metabolic Profiling of S-praziquantel: Structure Elucidation Using the Crystalline Sponge Method in Combination with Mass Spectrometry and Nuclear Magnetic Resonance.

Praziquantel (PZQ) is the drug of choice for treatment of the neglected tropical disease schistosomiasis. Although the drug has been extensively used over several decades and its metabolism well studied (several oxidative metabolites are known from literature), the knowledge of the complete structure of some of its metabolites remains elusive. Conventional techniques, such as nuclear magnetic resonance or liquid chromatography mass spectrometry were used in the past to investigate phase I and phase II metabolites of PZQ. These techniques are either limited to provide the complete molecular structure (liquid chromatography mass spectrometry) or require large amount of sample material (NMR), which are not always available when in vitro systems are used for investigation of the metabolites. In this study, we describe new structures of S-PZQ metabolites generated in vitro from human liver microsomes using the crystalline sponge method. After chromatographic separation and purification of the oxidative metabolites, ultra-performance liquid chromatography-quadrupole time-of-flight mass spectrometry analysis was conducted to narrow down the position of oxidation to a certain part of the molecule. To determine the exact position of hydroxylation, singe-crystal X-ray diffraction analysis of the crystalline sponges and absorbed analyte was used to identify the structure of S-PZQ and its metabolites. The crystalline sponge method allowed for complete structure elucidation of the known metabolites S-trans-4'-hydroxy-PZQ (M1), S-cis-4'-hydroxy-PZQ (M2) and S-/R-11b-hydroxy-PZQ (M6) as well as the unknown metabolites S-9-hydroxy-PZQ (M3) and S-7-hydroxy-S-PZQ (M4). For comparison of structural elucidation techniques, one metabolite (M3) was additionally analyzed using NMR. SIGNIFICANCE STATEMENT: The information content of the metabolic pathway of praziquantel is still limited. The crystalline sponge method allowed the complete structural elucidation of three known and two unknown metabolites of S-praziquantel, using only trace amounts of analyte material, as demonstrated in this study.

Crystalline Sponges as a Sensitive and Fast Method for Metabolite Identification: Application to Gemfibrozil and its Phase I and II Metabolites.

Understanding the metabolism of new drug candidates is important during drug discovery and development, as circulating metabolites may contribute to efficacy or cause safety issues. In the early phase of drug discovery, human in vitro systems are used to investigate human relevant metabolism. Though conventional techniques are limited in their ability to provide complete molecular structures of metabolites (liquid chromatography mass spectrometry) or require a larger amount of material not available from in vitro incubation (nuclear magnetic resonance), we here report for the first time the use of the crystalline sponge method to identify phase I and phase II metabolites generated from in vitro liver microsomes or S9 fractions. Gemfibrozil was used as a test compound. Metabolites generated from incubation with microsomes or S9 fractions, were fractionated using online fraction collection. After chromatographic purification and fractionation of the generated metabolites, single crystal X-ray diffraction of crystalline sponges was used to identify the structure of gemfibrozil metabolites. This technique allowed for complete structure elucidation of 5'-CH2OH gemfibrozil (M1), 4'-OH gemfibrozil (M2), 5'-COOH gemfibrozil (M3), and the acyl glucuronide of gemfibrozil, 1-O-ß-glucuronide (M4), the first acyl glucuronide available in the Cambridge Crystallographic Data Centre. Our study shows that when optimal soaking is possible, crystalline sponges technology is a sensitive (nanogram amount) and fast (few days) method that can be applied early in drug discovery to identify the structure of pure metabolites from in vitro incubations. SIGNIFICANCE STATEMENT: Complete structure elucidation of human metabolites plays a critical role in early drug discovery. Low amounts of material (nanogram) are only available at this stage and insufficient for nuclear magnetic resonance analysis. The crystalline sponge method has the potential to close this gap, as demonstrated in this study.

Synthesis and crystal structure of rac-2-(1,3-dioxo-isoindolin-2-yl)ethyl 4-methyl-N-phenyl-N'-(tri-iso-propyl-sil-yl)benzene-sulfondiimidoate: the first member of a new substance class.

The title compound {systematic name: rac-2-[7-methyl-4-(4-methylphenyl)-4-(phenylimino)-6,6-bis(propan-2-yl)-3-oxa-4λ6-thia-5-aza-6-silaoct-4-en-1-yl]-2,3-dihydro-1H-isoindole-1,3-dione}, C32H41N3O3SSi, was synthesized by desoxychlorination of 4-methyl-N-phenyl-N'-(triisopropyl-sil-yl)benzene-sul-fon-imid-am-ide and subsequent reaction with 2-(2-hy-droxy-eth-yl)isoindoline-1,3-dione. The racemic compound was crystallized from isopropanol. The structural characterization by single-crystal X-ray diffraction revealed two double-bonded nitro-gen atoms to the central sulfur atom and an overall crystal packing driven by its aromatic inter-actions.

Crystalline sponge affinity screening: A fast tool for soaking condition optimization without the need of X-ray diffraction analysis.

Structural elucidation of small molecules only available in low quantity (nanogram) is one of the big advantages of the crystalline sponge method. The optimization of various soaking parameters is crucial for effective analyte absorption and repetitive positioning in the pores of the crystal. Time-consuming X-ray diffraction measurements are necessary for data collection and confirmation of successful guest inclusion. In this work, we report a screening method to select optimal soaking conditions without the need of single-crystal X-ray diffraction analysis for individual compounds and mixtures. 14 substances were chosen as test compounds. Parallel guest soaking of individual compounds and mixtures was conducted using various soaking conditions. After evaporation of solvent, excessive material was removed, and guest molecules released through dissolution of the framework. Liquid chromatography-tandem mass spectrometry allowed the estimation of analyte trapped in the pores and the selection of optimal soaking condition dependent on the highest amount of analyte to crystal size (affinity factor). The tool allowed subsequent crystallographic analysis of ten compounds with minimal experiment time. Additionally, a study to examine the lower limit of detection of the crystalline sponge method was conducted. Determination of two target analytes was possible using only 5 ng of sample. Our study shows the potential of an affinity screening to prioritize soaking parameters by estimation of the guest concentration in a single crystal for one or multiple target compounds within a short period of time.