Preparation and chromatographic performance of cardanol-bonded silica stationary phase

doi:10.3724/SP.J.1123.2021.12023

Abstract

Abstract:

As green, less toxic, widely available, and site-rich functional ligands, natural products are widely used for the development of chromatographic stationary phases. In this work, a novel stationary phase, cardanol-bonded on silica (CBS) was prepared using γ-glycidoxypropyltrimethoxysilane (KH-560) as the coupling reagent and cardanol as the functional ligand. The synthesized stationary phase was characterized by Fourier transform-infrared spectra (FT-IR), thermogravimetric analysis (TGA), elemental analysis (EA), and N₂ adsorption-desorption analysis. The results revealed that cardanol was successfully immobilized on the surface of spherical silica by the ring-opening reaction of the epoxy groups in phenolic hydroxyl. The retention mechanism and chromatographic performance of the CBS column were further evaluated and compared with those of a commercial C₁₈ column using different classes of analytes, e. g., Tanaka standard test mixtures, alkylbenzenes, polycyclic aromatic hydrocarbons (PAHs), phenols, and aromatic positional isomers. The retention of alkylbenzenes under different chromatographic conditions revealed that the CBS column was a typical reversed-phase liquid chromatographic column, similar to the commercial C₁₈ column. From the results of the Tanaka test, it was concluded that CBS could provide various interactions for different solutes e. g., hydrogen bonding and π-π interactions, along with hydrophobic interactions. The synergistic effects resulting from the aromatic rings, the hydroxyl and alkyl linkers in the new stationary phase improved the separation selectivity via multiple retention mechanisms. Based on these interactions, different solute probes such as hydrophobic alkylbenzenes, PAHs, and phenols were successfully separated in the reversed-phase liquid chromatography (RPLC) mode. For example, the aromatic positional isomers o-terthenyl, m-terphenyl, and triphenylene were used to investigate the chromatographic performance of the CBS column. These PHAs were baseline separated with good peak shapes. The resolution of m-terphenyl and triphenylene was as high as 6.81, while the two isomers could not be separated on the C₁₈ column under the same chromatographic conditions. The repeatability and column stability of the CBS column was evaluated, and excellent repeatability and column stability were observed. The relative standard deviations (RSDs) of the retention time, peak area, and peak height for alkylbenzenes with 10 replicate injections were 0.052%-0.079%, 0.104%-0.847%, and 0.081%-0.272%, respectively. Traditional Chinese medicines have contributed notably to the Chinese civilization and human health. However, the complicated chemical compositions, unclear medicinal action mechanisms, and low purification efficiency for the traditional Chinese medicines have limited further development. Therefore it is necessary to establish an efficient, simple and feasible method for the separation and purification of herbal medicines. HPLC has been widely used in traditional Chinese medicines for the separation and detection of various components. In order to explore the CBS column for analysis of the traditional Chinese medicines, the ethanol extracts of fruits of Evodiae fructus and Camptotheca acuminata were used to test the separation performance of this column. The resolution of camptothecin from the preceding and following impurity peaks was 4.23 and 2.71. The resolution between evodiamine and rutaecarpin was 5.43, while the resolution from the adjacencies of impurity peaks was 2.20 and 1.69, respectively. The above mentioned results indicated that the CBS column shows good separation performance for the main active ingredients in the ethanolic extracts of these drugs, this validating its great potential for the analysis of real samples. Overall, the present study not only provides a new approach for the preparation of chromatographic stationary phases but also opens a new possibility for the separation and purification of camptothecin and evodiamine in real samples. This is an extension of the application of cardanol to chromatographic separation materials.

Key words: cardanol, bonded stationary phase, chromatographic properties, separation of traditional Chinese medicines

CLC Number:

O658

ZENG Lei, JIANG Lijuan, YAO Xingdong, WANG Ting, SHI Bo’an, LEI Fuhou. Preparation and chromatographic performance of cardanol-bonded silica stationary phase[J]. Chinese Journal of Chromatography, 2022, 40(6): 547-555.

Figures/Tables 15

Fig. 1 Synthesis of cardanol-bonded silica stationary phase

Table 1 Elemental analysis results for BS, KBS, and CBS

Analyte	Elemental contents/%
Analyte	C	H
BS	0.735	1.121
KBS	8.465	1.564
CBS	15.130	2.181

Table 2 BET surface area, pore volume, and pore size data

Analyte	BET surface area/ (m²/g)	Pore volume/ (cm³/g)	Pore size/ nm
BS	354.82	0.847	7.65
KBS	293.27	0.632	6.30
CBS	212.16	0.458	6.28

Fig. 2 FT-IR spectra of (a) BS, (b) KBS, and (c) CBS

Fig. 3 Thermogravimetric analysis curves of BS, KBS, and CBS

Fig. 4 Relationship between log k of alkylbenzenes and methanol volume fraction on CBS column Mobile phase: methanol/water; flow rate: 1.0 mL/min.

Fig. 5 Relationship between log k and alkyl chain length of alkylbenzenes on CBS and C18 columns Mobile phase: methanol/water (80∶20, v/v); flow rate: 1.0 mL/min.

Fig. 6 Radar plots of the Tanaka parameters of CBS and C18 columns kPB: retention factor of n-pentylbenzene, kn-pentylbenzene, used to evaluate the carbon content or density of ligands in the stationary phases; α C H 2: kn-pentylbenzene, used to evaluate the hydrophobicity of the stationary phase; αT/O : ktriphenylene/ko-terphenyl, used to evaluate the shape selectivity of the stationary phase; αC/P: kcaffeine/kphenol, used to evaluate the hydrogen-bonding interaction of the stationary phase; αB/P: kbenzenamine/kphenol, used to evaluate the apparent silanol activity of the stationary phase; αPB/O: kn-pentylbenzene/ko-terphenyl, used to describes the aromatic selectivity of the stationary phase.

Fig. 7 Chromatograms of Tanaka standard test mixture on CBS and C18 columns Mobile phase: methanol/water (75∶25, v/v); flow rate: 1.0 mL/min. Tanaka standard test mixture: 1. uracil; 2. benzenamine; 3. phenol; 4. caffeine; 5. n-butylbenzene; 6. n-pentylbenzene; 7. o-terphenyl; 8. triphenylene.

Fig. 8 Chromatograms of PAHs on CBS and C18 columns Mobile phase: methanol/water (80∶20, v/v); flow rate: 1.0 mL/min. Peaks: 1. benzene; 2. naphthalene; 3. biphenyl; 4. fluorene; 5. anthracene; 6. pyrene; 7. benz(a)anthracene.

Fig. 9 Relationships between log k and log P of solutes on CBS column and C18 columns Mobile phase and flow rate are the same as those in Fig. 8. Solutes: 1. benzene; 2. toluene; 3. ethylbenzene; 4. n-propylbenzene; 5. n-butylbenzene; 6. n-pentylbenzene; 7. naphthalene; 8. fluorine; 9. anthracene; 10. benz(a)anthracene.

Fig. 10 Chromatograms of aromatic positional isomers on CBS and C18 columns Mobile phase and flow rate are the same as those in Fig. 8.

Fig. 11 Chromatograms of phenols on CBS and C18 columns Mobile phase: methanol/water (60∶40, v/v); flow rate: 0.8 mL/min. Peaks: 1. phloroglucinol; 2. resorcinol; 3. phenol; 4. 2-cresol; 5. 3-nitrophenol; 6. 2-naphthol.

Fig. 12 Separation repeatability of the CBS column Mobile phase: methanol/water (80∶20, v/v); flow rate: 0.8 mL/min.

Fig. 13 Chromatograms of (a) Camptotheca acuminate extract and (b) Fructus evodiae extract on the CBS column

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