Saliramophenol, an unprecedented natural t-butylphenol derivative from Salicornia ramosissima J. Woods

Abstract

Saliramophenol (1), a unique skeleton-type compound with four t-butyl groups, was isolated from aerial parts of S. ramosissima J. Woods. The structure of the isolated compound was elucidated by detailed NMR spectroscopic methods and mass spectrometry data.

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The interest in the Salicornia species (Chenopodiaceae family) is growing due its versatile commercial products with great nutritional value,^1–3 along with its ethnopharmacological applications.³ Simultaneously Salicornia species are a source of bioactive metabolites.³ Salicornia ramosissima J. Woods is an annual halophyte usually erect, 3–40 cm tall, fleshy, with stem articulated and branched apparently leafless,⁴ included in the species aggregate S. europaea agg. and that grows naturally on the salt marsh of Europe.⁵ From the chemical profile point just recently its lipophilic profile was characterized⁶ and some of its phenolic constituents were identified.⁷ As a part of our continuing search for structurally interesting natural products from plants, the dichloromethane extract from aerial parts of S. ramosissima was phytochemically investigated and as a result a new natural t-butylphenol derivative, named saliramophenol (1) (Fig. 1) was isolated from this halophyte collected in Marina dos Puxadoiros (40°39′23″ N, 8°40′35″ W), Ria de Aveiro, Portugal (yield 0.00055% of dried plant). Herein, the isolation and structure elucidation of 1 is described.

Fig. 1 Chemical structure of saliramophenol (1).

Compound (1) 2,2′-((oxybis(5-(tert-butyl)-2,1-phenylene))bis(propane-2,2-diyl))bis(4-(tert-butyl)phenol), named saliramophenol, is a new natural phenol derivative with four tert-butyl groups. It was isolated as a pale yellow oil and has the molecular formula of C₄₆H₆₂O₃, which was established on the basis of its HRMS-ESI(+) peak at m/z 663.4773 [M + H]⁺ (calcd for C₄₆H₆₃O₃, 663.4772). Additionally, MS/MS analysis showed consecutive losses of m/z = 56 from the signal with m/z = 663 ([M + H]⁺), which are consistent with the loss of the four t-butyl groups. Another piece of structural evidence is shown by further MS/MS analysis of the signal with m/z = 685 ([M + Na]⁺). In this case not only is the loss of the t-butyl groups evidenced but a signal with m/z = 251 is also observed which corresponds to the mass of a fragment with two aromatic rings, thus indicating an ether linkage between carbons C-1′ and C-1″.

The ¹³C NMR data (Table 1) and the DEPT spectrum showed only four non-equivalent sp³ carbons (two methylic and two quaternary carbons) and eight non-equivalent sp² carbons (three CH and five deprotonated aromatic carbons). These features reveal a high degree of symmetry in the chemical structure of compound (1). The compound also possesses two quasi-equivalent 1,2,4-trisubstituted aromatic rings, as evidenced by the presence of the downfield ¹H NMR signals at δ_H 7.13, as a double doublet (J = 2.5 and 8.6 Hz), at δ_H 7.53 as a doublet (J = 8.6 Hz) and at δ_H 7.35 and 7.36 as two doublets (J = 2.5 Hz), the coupling between these protons was confirmed by the COSY NMR spectrum.

Table 1 ¹H and ¹³C NMR chemical shifts for saliramophenol (1) in CDCl₃

Position	Compound 1
	δH (J in Hz)	δC, type
a Those marked with the same symbol may be exchanged.b Those marked with the same symbol may be exchanged.c Those marked with the same symbol may be exchanged.
1		147.6, C^a
2		138.5, C^b
3	7.35 (1H, d, 2.5)^c	120.4, CH
4		147.0, C
5	7.13 (1H, dd, 2.5, 8.6)	124.0, CH
6	7.53 (1H, d, 8.6)	119.1, CH
7		34.5, C
8	1.28 (9H, s)	31.4, CH3
9		34.8, C
10	1.33 (6H, s)	30.2, CH3
1′		147.7, C^a
2′		138.4, C^b
3′	7.36 (1H, d, 2.5)^c	120.4 CH
4′		147.0, C
5′	7.13 (1H, dd, 2.5, 8.6)	124.0, CH
6′	7.53 (1H, d, 8.6)	119.1, CH
7′		34.5, C
8′	1.28 (9H, s)	31.4, CH3
9′		34.8, C
10′	1.33 (6H, s)	30.2, CH3
1″		147.7, C^a
2″		138.4, C^b
3″	7.36 (1H, d, 2.5)^c	120.4, CH
4″		147.0, C
5″	7.13 (1H, dd, 2.5, 8.6)	124.0, CH
6″	7.53 (1H, d, 8.6)	119.1, CH
7″		34.5, C
8″	1.28 (9H, s)	31.4, CH3
1‴		147.6, C^a
2‴		138.5, C^b
3‴	7.35 (1H, d, 2.5)^c	120.4, CH
4‴		147.0, C
5‴	7.13 (1H, dd, 2.5, 8.6)	124.0, CH
6‴	7.53 (1H, d, 8.6)	119.1, CH
7‴		34.5, C
8‴	1.28 (9H, s)	31.4, CH3

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The presence of four equivalent t-butyl groups attached to the aromatic rings was deduced from the following spectroscopic evidence: (i) a singlet at δ_H 1.28 (integral proportional to 36 protons) showing ¹J_H–C correlation (HSQC spectrum) with the signal at δ_C 31.4, corresponding to nine methyl groups (Table 1); (ii) this ¹H NMR signal showed connectivities (HMBC spectrum) with the signals at δ_C 31.4, 34.5 and 147.0 (Fig. 2a), corresponding, respectively, to the resonance of the methylic carbons (C-8, C-8′, C-8″ and C-8‴), aliphatic quaternary carbons (C-7, C-7′, C-7″ and C-7‴) and deprotonated aromatic carbons (C-4, C-4′, C-4″ and C-4‴).

Fig. 2 Coupling deduced by the COSY spectrum (bold line) and HMBC connectivities (→), observed for compound 1.

The ¹H NMR spectrum also showed a singlet at δ_H 1.33, corresponding to the resonance of the protons from 4 equivalent methyl groups (δ_C 30.2), and that exhibits HMBC connectivities with the ¹³C NMR signals at δ_C 30.2, δ_C 34.8 (aliphatic quaternary carbon) and δ_C 138.4, 138.5 (Fig. 2b), showing that the two t-butylphenol units are linked by an isopropyl-type group.

The most downfield ¹³C NMR signals (δ_C 147.6 and 147.7) correspond to two non-equivalent oxygen-bearing carbons in position C-1, C-1‴ and C-1′, C-1″. The FTIR absorption band at ν_max 3443 cm⁻¹ suggests the presence of an OH group attached to an aromatic moiety.

All these spectroscopic data and other HMBC connectivities (Fig. 2a and b) are only compatible with the structure presented here for compound 1, saliramophenol (Fig. 1).

Natural compounds with t-butyl groups are described in the literature^8,9 and t-butylphenol is a known commercial product used in the synthesis of resins so it can be widespread in the environment.^10,11 Saliramophenol 1, a new natural compound with a unique carbon skeleton, which is reported here, cannot be an artifact since the bond between the aromatic rings cannot be formed during the isolation and/or purification procedures, so it must be biosynthesized using t-butylphenol as substrate and with the intervention of enzymes. It seems that saliramophenol 1 could be one of the first examples of compounds isolated from plants that are a result of biogenetic pathways using anthropogenic substrates.

There are examples of biologically active compounds possessing t-butyl phenol units, for example succinobucol is a synthetic antiatherosclerotic drug proved to have antihyperglycemic activity and is in phase III clinical trials.¹² There is also evidence that the t-butyl group seems to play an essential role in the platelet activating factor receptor antagonist activity.¹³ This unique structure in a natural compound is most likely due to the environmental stress of the S. ramosissima habitat (e.g. salinity, water supply).

In summary, saliramophenol 1 is the first example of a natural compound with a dimeric structure of two t-butylphenol units linked by an isopropyl-type carbon–carbon skeleton. The isolation and unequivocal structural elucidation of 1 has added a completely new skeleton to the already large and varied family of chemical structures obtained from natural resources. Further investigations regarding the biological activity of compound 1 are in progress.

Acknowledgements

Thanks are due to FCT/MEC for the financial support to the QOPNA research Unit (FCT UID/QUI/00062/2013), through national funds and where applicable co-financed by the FEDER, within the PT2020 Partnership Agreement and to the University of Azores, Centre for Environmental and Marine Studies (CESAM) and the Portuguese National NMR Network (RNRMN).

Notes and references

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Footnote

† Electronic supplementary information (ESI) available: 1D and 2D NMR, MS and HRMS spectra of saliramophenol (1) and detailed experimental procedure. See DOI: 10.1039/c5ra10893d

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