Structure Elucidation Using Strict Structure Generation

Actinomycetes are well recognized as the richest source of bioactive compounds, including clinically important antibiotics, antitumor agents and cell function modulators, and hence of high pharmacological and commercial interest.1 Amongst this group, members of the genus Streptomyces are the most prolific producers of secondary metabolites, accounting for up to 80% of the bioactive small molecules discovered from actinomycetes.2 Meanwhile, it is quite notable that further discovery of unknown metabolites from Streptomyces is predicted by the genome analysis: the number of metabolites actually isolated is far more below the number of secondary metabolite biosynthetic gene clusters identified in the whole genomes of S. avermitilis and S. coelicolor.3,4 As a part of our chemical investigation on microbial secondary metabolites, we reported plant-growth promoting spiroacetals of polyketide origin,5 a linear polyketide with a dlactone terminus with cytotoxic activity,6 a polycyclic tetronate with antiinvasive activity7 and a biosynthetically unprecedented heterocyclic polyketide with antibacterial and antiinvasive activities8 from Streptomyces. During the course of our continuing effort to discover structurally unique secondary metabolites from these organisms, a new modified peptide was isolated from the culture broth of a soilderived actinomycete strain Streptomyces sp. SPMA113 collected in Thailand. The strain was cultured in A-11M liquid medium, and the whole culture broth was extracted with 1-butanol. The HPLC/UV analysis of the extract using our in-house metabolite database indicated the presence of an unknown compound showing a UV absorption maximum at 260nm, along with geldanamycins9 and elaiophylins.10 Guided by HPLC/UV, several steps of chromatographic purification resulted in the isolation of a new compound, prajinamide (1, Figure 1). Compound 1 was obtained as a pale yellow oil that analyzed for a molecular formula of C16H25N3O3 (6 degrees of unsaturation) by interpretation of HR–ESI–TOF–MS (observed [M+Na]+ at m/z 330.1788, calculated [M+Na]+ 330.1788). This molecular formula was corroborated by 1H and 13C NMR spectral data (Table 1). Analysis of the combined 1D and 2D NMR data established that 1 possessed three carbonyl, four olefinic methine, two sp3 methine, five sp3 methylene and two methyl carbons, in addition to three exchangeable protons. The IR absorptions at 1647, 1599 and 1538 cm 1 indicated the presence of amide functionalities, which was supported by the resonances of carbonyl carbons at d 165.6, 169.9 and 170.1 observed in the 13C NMR spectrum. As three carbonyls and two double bonds accounted for five of six double-bond equivalents, 1 must be a monocyclic compound. Further analysis of 1H-1H COSYand HMBC spectra provided three substructures (Figure 2). The first was an ornithine lactam that was established by the sequential COSY correlations from an NH proton at d 7.61 (2-NH) to another NH proton at d 8.07 (5-NH) through a methine proton (H-2) and three methylene protons (H-3, H-4 and H-5) and HMBC correlations from H-2, H-3, H-5 and 2-NH to a carbonyl carbon C-1 (d 169.9). The second substructure, a b-alanine, was assigned on the basis of COSY correlations between an NH proton at d 7.98 (8-NH) and H-8 and between H-8 and H-7 and HMBC correlations from H-7 and H-8 to C-6 (d 170.1). Two and three-bond C–H correlations from 2-NH and H-2 to C-6 allowed the b-alanine residue being connected to the ornithine lactam through an amide linkage. COSY correlations between four olefinic protons H-10, H-11, H-12 and H-13 provided a conjugated diene, which was then extended to include a carbonyl carbon C-9 (d 165.6) at C-10 on the basis of HMBC correlations from H-10 and H-11 to C-9. A vinyl methine H-13 showed a COSY correlation to H-14, which showed in turn correlations to two equivalent methyl protons H-15 and H-16, thereby establishing an isopropyl terminus attached to the diene fragment. The geometries of C-10–C-11 and C-12–C-13 double bonds were assigned as Z and E, respectively, on the basis of the vicinal coupling constants (JH10,H111⁄411.3Hz, JH12,H131⁄415.5Hz). The third substructure was thus

Expert systems for spectroscopic molecular structure elucidation have been developed since the mid-1960s. Algorithms associated with the structure generation process within these systems are deterministic; that is, they are based on graph theory and combinatorial analysis. A series of expert systems utilizing 2D NMR spectra have been described in the literature and are capable of determining the molecular structures of large organic molecules including complex natural products. Recently, an opinion was expressed in the literature that these systems would fail when elucidating structures containing more than 30 heavy atoms. A suggestion was put forward that stochastic algorithms for structure generation would be necessary to overcome this shortcoming. In this article, we describe a comprehensive investigation of the capabilities of the deterministic expert system Structure Elucidator. The results of performing the structure elucidation of 250 complex natural products with this program were studied and generalized. The conclusion is that 2D NMR deterministic expert systems are certainly capable of elucidating large structures (up to about 100 heavy atoms) and can deal with the complexities associated with both poor and contradictory spectral data.

The fundamental operational modes of 1D CASE systems provide insight into the approaches used to overcome many of the difficulties encountered in the development of efficient expert systems, including modern 2D CASE systems. In this Chapter we will focus on 1D NMR based expert systems developed from the end of the 1990s to the present. We will discuss how researchers have directed their efforts to improve the methods for the extraction of structural information from spectra, identifying optimal strategies for computer-assisted structure elucidation and increasing the user-friendliness of the systems. The strategies include using knowledge bases, structural-group analysis and structure generation and verification. We will review the implementation of these strategies in a number of systems including X-PERT, SpecSolv and Structure Elucidator. We will also discuss the limitations of 1D CASE systems in terms of the size and complexity of the molecules that can be studied and conclude that complex chemical structures can only be elucidated when both 1D and 2D NMR spectral data are used.

One new triterpenoid, 3β, 16β, 23, 28-tetrahydroxy oleana-11, 13(18)-dien-3-O-β-D-glucopyranosyl(1 → 3) [β-D-glucopyranosyl(1 → 2)]-β-D-fucopyranoside (1), together with two known flavonoids, were isolated from Stellaria media (L.) Cyr. The structure elucidation of the new compound was primarily based on HREIMS, EIMS, UV, IR, 1D- and 2D-NMR analyses, including COSY, HMQC and HMBC correlations.

Phytochemical investigation of Platytaenia multicaule resulted in the isolation of two new ceramides, N-triacontylpentadecanamide (1) and 1,3,4-trihydroxy-2,dodecanoylamino-(24E)-unacotenene (2), along with two known compounds, β-sitosterol (3) and β-sitosterol glycoside (4). The structure elucidation of the isolated compounds was based primarily on 1D and 2D NMR analyses, including COSY, NOESY, HMQC, and HMBC correlations.

Computer-assisted structure elucidation (CASE) is composed of two steps: (a) generation of all possible structural isomers for a given molecular formula and 2D NMR data (COSY, HSQC, and HMBC) and (b) selection of the correct isomer based on empirical chemical shift predictions. This method has been very successful in solving structural problems of small organic molecules and natural products. However, CASE applications are generally limited to structural isomer problems and can sometimes be inconclusive due to insufficient accuracy of empirical shift predictions. Here, we report a synergistic combination of a CASE algorithm and density functional theory calculations that broadens the range of amenable structural problems to encompass proton-deficient molecules, molecules with heavy elements (e.g., halogens), conformationally flexible molecules, and configurational isomers.

Phytochemical investigations of the whole plant of Stachys parviflora (Lamiaceae) resulted in the isolation of two new phenethyl alcohol glycosides. The structures of the new compounds named parviflorosides A and B were established as 2-(3,4-dihydroxyphenyl)-ethyl-O-α-l-rhamnopyranosyl-(1 → 2)-4-O-E-caffeoyl-β-d-glucopyranoside (1) and 2-(3,4-dihydroxyphenyl)-ethyl-O-α-l-rhamnopyranosyl-(1 → 2)-6-O-E-caffeoyl-β-d-glucopyranoside (2), respectively. The structure elucidation of the new compounds was based primarily on 1D and 2D NMR analysis, including COSY, HMBC and HMQC correlations.

Two new iridoids 1,3-dimethyloxy-7-hydroxymethyl-4-(3-methyl-butyryloxymethyl)-1-hydrocyclopenta-4,7-diene[c]pyran-6-one (1) and 1,3-dimethyloxy-7-hydroxymethyl-4-methyloxymethyl-1-hydrocyclopenta-4,7-diene[c]pyran-6-one (2) were isolated from the roots of Patrinia scabra Bunge. The structure elucidation of the isolated compounds was based primarily on HRESIMS, EIMS, IR, UV, 1D- and 2D-NMR analyses, including COSY, HMQC, HMBC and NOESY correlations, as well as X-ray crystallographic analysis.

Four new pterocarpans, atricarpan A (=(-)-1,2-dihydroxy-4-(hydroxymethyl)-3,9-dimethoxypterocarpan; 1), atricarpan B (=(-)-2,3-ethylenedioxy)-1,4-dihydroxy-9-methoxypterocarpan; 2), atricarpan C (=(-)-1,9-dimethoxypterocarpan-3-carboxylic acid; 3), and atricarpan D (=(-)-2,9-dimethoxy-4-(5-oxohexyl)pterocarpan; 4) were isolated from the BuOH extract of the whole plant of Zygophyllum eurypterum. The structure elucidations of those compounds were based primarily on 1D- and 2D-NMR analysis, including COSY, HMBC, and HMQC correlations. Compounds 1-4 also inhibited butyrylcholinesterase (BChE; EC 3.1.1.8) enzyme in a concentration-dependent manner with IC(50) values between 12.5-65.0 microM. Similarly, compounds 1 and 4 inhibited lipoxygenase (LOX; EC 1.13.11.12) and acetylcholinesterase (AChE; EC 3.1.1.7) enzymes with IC50 values of 13.5 and 20.5 muM, respectively.

Nuclear magnetic resonance (NMR) is a powerful technique for the structure elucidation of organic molecules and significantly important in the pharmaceutical industry supporting the discovery and development of drug substances or active pharmaceutical ingredients (APIs). Following an initial study and assessment of a prototype NMR instrument with a high-temperature superconducting (HTS) power-driven magnet of 9.4 T (400 MHz for 1H observation) operating with standard commercial electronics and probes, we tested the instrument with three compounds representing typical pharmaceutical drugs. We compared results from two probes and shimstacks with different geometries (broadband fluorine observe (BBFO) with Bruker orthogonal shim system-3 (BOSS3) and Bruker quattro nucleus probe (QNP) with BOSS1 shims) testing standard one-dimensional (1D) NMR experiments including selective excitation experiments, and two-dimensional (2D) homonuclear and heteronuclear experiments for the purposes of evaluating the equipment for structure elucidation capabilities. In our initial study on cinacalcet HCl, only the 1D 1H experiment showed a loss of resolution when using the longer coiled BBFO probe with the BOSS3 shims compared to the shorter coiled QNP probe with BOSS1 shims. The selective excitation experiments using the cinacalcet HCl were successful. Our characterization of 1D (1H, 13C, 19F) and 2D (1H-1H and 1H-13C) NMR experiments for compounds I and II with the two probes and shimstacks indicated no significance differences. Overall, these results are satisfactory with the HTS magnet at the field of 9.4 T for the structural elucidation work of standard pharmaceutical compounds. This new technology has the advantage of being able to locate the HTS NMR magnet system in any chemistry or analytical laboratory where the samples are produced, facilitating rapid analysis with minor needs from the facilities and without cryogenic liquids.

A new cucurbitacin, endecaphyllacin C, was isolated from the tubers of Hemsleya endecaphylla. The structure was elucidated as 2β,16α,20β,25-tetrahydroxy-24-acetylaminocucurbita-5-en-3,11,22-trione (1) on the basis of extensive 1D and 2D NMR techniques, including COSY, HMBC, HMQC, and NOESY correlations, as well as HR-FAB-MS analysis.

Case, a computer model of the structure elucidation process

Purely on the basis of his scientific achievements, Carl Djerassi must be considered one of the preeminent organic chemists of the 20th century. In a career that has spanned almost five decades, Djerassi has been active in the development of new medicinal agents, including the synthesis of the first active oral contraceptive; the isolation, structure elucidation, and partial synthesis of several hundred natural products; and investigations into aspects of the biological function and biosynthesis of those natural products. Perhaps most important scientifically, Djerassi has played a central role in developing several physical methods for the solution of stereochemical and structural problems in chemistry. His research has led to the publication of more than 1100 papers and seven books, which he either authored or coauthored. This year, the American Chemical Society awarded him its highest honor, the Priestley Medal, in recognition of his accomplishments. Although many distinguished chemists make important ...

In the real world problems, mostly there is difference between dimensions of simulated and corresponding fabricated structures. In case of Frequency Selective Surface (FSS), this variation can be even larger as the frequency gets into terahertz range. One of the approach to study such variations is metamodeling. In this work, we address the problem of modeling such structures with statistical variations in their geometries by using artificial neural network (ANN). We train and test this metamodel and finally show its performance statistics.

Design of Mn(II), Fe(III) and Ru(III) chalcone complexes: Structural elucidation, spectral, thermal and catalytic activity studies