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Piperkadsin C Structure Elucidation

October 31, 2022

Piperkadsin C

This example application of ACD/Structure Elucidator (ACD/SE) comes from a publication by M. Elyashberg and coworkers [1] devoted to structure revisions of a set of natural products whose structures were already published in literature.

Kim and coworkers [2] isolated a new neolignan, piperkadsin C (1), and elucidated its structure from HRMS and NMR spectra (1H, 13C, HSQC, HMBC).

1

Compound 1 was obtained as a viscus gum. In the positive mode FABMS of 1, a quasimolecular ion peak at 340 [M]+ was observed, and the molecular formula of 1 was determined to be C20H20O5 by HR-FABMS: observed, 340.1315 (calculated for C20H20O5, 340.1311).

However, this structure was revised by the same group in their next article [3]. In the positive mode FABMS, a molecular ion peak [M+H]+ at m/z 357 was observed, and the molecular formula of the compound was determined to be C20H20O6 by the HR-FABMS spectra and a molecular ion peak [M+H]+ at m/z 357.1335 was observed (calcd. for C20H20O6, 357.1338). The following revised structure was proposed:

2

As we see, the molecular formula of the revised structure contains one additional oxygen. It is interesting that in both cases calculated and measured molecular masses agreed with great accuracy.

To verify the revised structure 2, the NMR spectroscopic data presented in the article [3] were entered into ACD/Structure Elucidator (see Table 1).

Table 1. NMR spectroscopic data of the structure 2

Label δC δC Calc (HOSE) CHn δH C to H HMBC
C 1 133.900 130.380 C
C 2 108.800 106.580 CH 6.920 C 7, C 6, C 1, C 4, C 3
C 3 147.600 147.410 C
C 4 146.800 147.170 C
C 5 108.100 107.780 CH 6.760 C 6, C 1, C 4, C 3
C 6 122.700 123.980 CH 6.750 C 7, C 5, C 2, C 1, C 4
C 7 65.300 68.450 CH 3.640 C 9, C 8, C 14, C 2, C
6, C 1, C 13, C 15
C 8 50.400 61.320 C
C 9 16.800 20.600 CH3 0.960 C 8, C 14, C 7, C 15
C 10 132.600 134.070 C
C 11 157.200 118.150 CH 7.000 C 16, C 12, C 10, C 13,
C 15
C 12 131.700 142.070 C
C 13 147.000 146.650 C
C 14 58.600 56.020 CH 3.360 C 9, C 8, C 7, C 12, C
10, C 13, C 15
C 15 197.200 191.390 C
C 16 33.000 35.700 CH2 2.910 C 18, C 10, C 17, C 11,
C 15
C 17 135.500 134.430 CH 5.820 C 16, C 18, C 10
C 18 116.700 116.990 CH2 5.080
C 18 116.700 116.990 CH2 5.060 C 16, C 10, C 17
C 19 101.100 101.220 CH2 5.940 C 4, C 3
C 20 59.200 55.130 CH3 3.820 C 12

The 13C chemical shifts assigned by authors [3] were set to carbon atoms of 2, and chemical shift prediction was performed using three algorithms implemented into the ACD/Structure Elucidator: HOSE code based, incremental and neural networks. Results are shown in Figure 1.

Figure 1. Structure of compound 2 proposed in [2] for which 13C chemical shift prediction was carried out using the HOSE code-based method, the neural networks, and the incremental approach. Average deviations of 13C chemical shifts determined by these methods are denoted as dA, dN and dI correspondingly. Each atom is colored to mark a difference between its experimental and calculated 13C chemical shifts. The green color represents a difference between 0 to 3 ppm, yellow was >3 to 15 ppm, red > 15 ppm. Red arrows indicate nonstandard HMBC correlations (NSC) – those whose lengths exceeds three chemical bonds (nJCH, n>3).
We see that the average deviations are large, which hinted that the structure is problematic. Figure 1 shows that the proposed structure contains four nonstandard correlations (NSC) of 4-bonds length. To verify the structure using ACD/SE, a Molecular Connectivity Diagram (MCD) was created in which the “green fragment” (confirmed by 13C prediction) was pre-defined (Figure 2). Also, the caron atom at 197.2 ppm was pre-defined to be a carbonyl, as there isn’t any other option for that chemical shift value.

Figure 2. Molecular connectivity diagram (MCD) created for revised structure 2. Hybridizations of carbon atoms are marked by corresponding colors: sp2 – violet, sp3 – blue. Labels “ob” and “fb” are set by the program to carbon atoms for which neighboring with heteroatom is either obligatory (ob) or forbidden (fb).  HMBC connectivities are marked by green arrows.

As structure 2 contains four NSCs, Fuzzy Structure Generation was run from the MCD (Figure 2) with the following options: the presence of four NSCs of unknown lengths was admitted; 13C chemical shifts were calculated during the generation; structural filtering was switched off to provide saving of the proposed structure 2. Results: k =667→(removal of duplicates)→606, tg = 44 s.

After 13C chemical shift prediction using all three methods, the output file was ranked in increasing order of dA average deviations. The  nine top ranked structures are presented in Figure 3.

Figure 3. The nine top ranked structures.

Figure 3 shows that all structures are characterized by large average and maximum deviations, while the proposed structure 2 was placed in 9th position by the ranking procedure. This means that all generated structures, including the revised one, are incorrect. We hypothesized that there is a probability to deduce a correct structure from the molecular formula and NMR data used for deducing structure 1.

With this in mind, we entered the indicated data (Table 2) into the ACD/SE and, as in the previous case, verified structure 1 (see Figure 4).

Table 2. NMR spectroscopic data used for deducing structure 1 [2]

Label δC δC calc (HOSE) CHn δH H to C HMBC
C 1 133.900 133.350 C
C 2 108.800 107.290 CH 6.920 C 7, C 6, C 1, C 4, C 3
C 3 147.600 148.960 C
C 4 146.800 147.570 C
C 5 108.100 108.010 CH 6.760 C 6, C 1, C 4, C 3
C 6 122.700 122.240 CH 6.750 C 7, C 5, C 2, C 1, C 4
C 7 65.300 75.920 CH 3.640 C 9, C 8, C 14, C 2, C
6, C 12, C 1, C 15
C 8 50.400 49.090 C
C 9 16.800 21.680 CH3 0.960 C 8, C 14, C 7, C 12, C
13, C 11
C 10 132.600 129.360 C
C 11 157.200 132.440 CH 7.000 C 16, C 8, C 12, C 10,
C 13, C 15
C 12 131.700 121.040 C
C 13 147.000 160.830 C
C 14 58.600 52.450 CH 3.360 C 9, C 8, C 7, C 12, C
10, C 13, C 15
C 15 197.200 197.350 C
C 16 33.000 35.440 CH2 2.910 C 18, C 10, C 17, C 11,
C 15
C 17 135.500 134.660 CH 5.820 C 16, C 18, C 10
C 18 116.700 116.910 CH2 5.080
C 18 116.700 116.910 CH2 5.060 C 16, C 10, C 17
C 19 101.100 101.270 CH2 5.940 C 4, C 3
C 20 59.200 59.030 CH3 3.820 C 13
Figure 4. Verification of structure 1.

Figure 4 shows that structure 1 is wrong, as it has large average and maximum deviations. Moreover, this structure contradicts Bredt’s rule and cannot exist due to its instability [1].

The MCD created from the data presented in Table 2 is shown below in Figure 5

Figure 5. MCD created from the data used for deducing structure 1.

Fuzzy Structure Generation was initiated with options selected automatically. Structure filtering was switched off as in the previous case. Results: k = 6, tg = 6 m 50 s, 4 (from 44) connectivities have been extended during generation. The ranked output file is shown in Figure 6.

Figure 6. The ranked output file.

We see that the two best structures #1 and #2 are similar and differ only by the positions of the OH and O-CH3 groups. The original structure 1 was placed in 5th position. We subsequently used the methodology suggested by authors of ref. [4]. On the basis of DFT-based 13C chemical shift prediction using the DU8+ approach, it was determined that structure #2 is the correct one with a Mean Average Error of 1.08 ppm. Structure #1 has a MAE value of 2.17 ppm.

Thus, the correct structure of piperkadsin C was determined as a result of the combined application of CASE and DFT-based chemical shift prediction for the top structures of the ranked output file.

References

  1. Elyashberg, M.; Novitskiy, I. M.; Bates, R. W.; Kutateladze, A. G.; Williams, C. M. (2022). Reassignment of Improbable Natural Products Identified through Chemical Principle Screening. European Journal of Organic Chemistry, e202200572. https://doi.org/10.1002/ejoc.202200572
  2. K.H. Kim, J. W.Choi, S. K. Ha, S.Y. Kim, K. R. Lee. (2010). Neolignans from Piper kadsura and their anti-neuroinflammatory activity. Bioorganic & Medicinal Chemistry Letters, 20, 409–412.
  3. K.H. Kim, J. W.Choi, S. K. Ha, S.Y. Kim, K. R. Lee. (2010). Corrigendum to ‘‘Neolignans from Piper kadsura and their anti-neuroinflammatory activity”. Bioorganic & Medicinal Chemistry Letters, 20, 3186–3187.
  4. A. V. Buevich, M. E. Elyashberg. (2016). J. Nat. Prod., 79 (12), 3105–3116.

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