Elsevier

Talanta

Volume 77, Issue 4, 15 February 2009, Pages 1299-1303
Talanta

Quantitative determination of chlorogenic acid in Honeysuckle using microwave-assisted extraction followed by nano-LC-ESI mass spectrometry

https://doi.org/10.1016/j.talanta.2008.09.003Get rights and content

Abstract

The chlorogenic acid (CA) in Honeysuckle is determined and identified by nano-liquid chromatography-electrospray ionization mass spectrometry (nano-LC-ESI/MS) after extraction with microwave-assisted extraction (MAE). As a new sample preparation method for Honeysuckle, the MAE procedure is optimized, validated and compared with conventional methods including reflux extraction (RE) and ultrasonic extraction (USE). It is found that MAE gives the best result due to the highest extraction efficiency within shortest extraction time (only 4 min). Here, CA is determined by nano-LC-ESI/MS based on the calibration curve of its authentic standard. The method linearity, detection limit, precision and recovery are studied. The results show that the combined MAE and nano-LC-ESI/MS method has a linearity (R2 0.991, 0.8–20 ng mL−1), a low limit of detection (0.5 ng mL−1), good precision (R.S.D. = 2.54%) and a recovery (84.8%). The experiment has demonstrated that the nano-LC-ESI/MS following MAE is a fast and reliable method for quantitative analysis of CA in Honeysuckle.

Introduction

Honeysuckle, the dried flower of Lonicera japonica Thunb., is a common Traditional Chinese Medicine (TCM). It has shown a wide spectrum of biological and pharmacological activities such as antibacterial, antiviral [1], antioxidant [2]. Honeysuckle contains chlorogenic acid (CA), luteolin-7-O-glucoside, volatile oil, flavone, saponins, polysaccharides, and polyphenolic compound [3]. CA is the major bioactive constituent in the herb. It is necessary to measure CA in Honeysuckle for quality of Honeysuckle.

Prior to the analysis, the extraction of CA in Honeysuckle is required. Reflux extraction (RE) is the most widely used traditional technique for routine herb drug extraction [4], [5], [6]. However, RE is laborious, time-consuming, and requires large amounts of solvents. Ultrasonic extraction (USE) is also applied for the CA extraction [7]. However, it does not show satisfactory extraction efficiency. Therefore, a fast, solvent-free and low-cost sample preparation technique for the extraction of CA from TCMs is desirable. Microwave heating involves internal heating based on conduction and dielectric polarization caused by microwave irradiation [8]. Recently, microwave-assisted extraction (MAE) has received more and more attention as a potential alternative to traditional solid–liquid extraction methods, mainly due to considerable savings in processing time and solvent consumption [9], [10]. It is more efficient when compared with traditional heating-reflux since microwave irradiation accelerates cell rupture by sudden temperature rise and internal pressure increase inside the cells of plant sample, which promotes destroy of sample surface [11]. It has been employed for the extraction of various biologically active compounds from different plant matrices [12], [13], [14], [15], [16], [17]. In our previous study [18], MAE followed by gas chromatography–mass (GC–MS) has been applied to the TCM analysis.

Recently, remarkable advances in high performance liquid chromatography (HPLC) have been made for the separation and analysis of components in TCMs. Nano-LC as a new powerful analytical tool is carried out in capillaries of small internal diameter (I.D. < 300 μm) containing stationary phases usually employed in HPLC. The nano-LC offers several advantages over classical analytical methods, e.g., rapid separation achieved in a short analysis time with high efficiency, use of minute volumes of mobile phases as well as small amounts of packing materials [19], [20], and low sample dilution ratio. However, the method does not prove to be satisfactorily sensitive due to the small volumes of sample injected and the short path length of the detection cell used in UV detection. The combination of nano-LC with mass spectrometry results in a powerful tool increasing the method sensitivity. ESI has now become the most important ionization technique for the on-line coupling of liquid phase separation techniques such as LC with MS [21], [22], [23]. Advantages of the coupling of LC and ESI/MS are that (a) ESI is the technique of choice for transferring the charged species from liquid phase into gas phase, (b) the hyphenated technique has a broad range of applications from ions and small molecules to macromolecules and (c) ESI-MS allows the detection of multiple charged species originating from high molecular mass molecules. The coupling of both techniques (nano-LC–MS) is easily achieved because of the relatively low flow rate, typical feature of nano-LC system, originated by the employment of small I.D. columns. Thus, nano-LC–MS is often used for the analysis of proteomics [24], pharmaceutical compounds [25] due to its high sensitivity and low limit of detection.

In this study, a MAE followed by nano-LC-ESI/MS is developed for the determination of CA from Honeysuckle. This is the first report of hyphenating MAE with nano-LC–MS. The MAE parameters are optimized, and the extraction efficiency by MAE is compared with that by conventional extraction techniques. The method linearity, recovery, precision and detection limit are also studied.

Section snippets

Chemical and materials

Methanol, acetonitrile and formic acid were chromatographic grade (Merk, Darmstadt, Germany). Ethanol was analytical grade. Distilled water was purified by a Milli-Q system (Milford, MA, USA). Honeysuckle was purchased from local Chinese medicine stores in Shanghai, China. Chlorogenic acid (purity 99.9%) was purchased from the Institute for the Control of Pharmaceutical and Biological Products of China (Beijing, China). Centrifuge was from Christ (Osterode am Harz, Germany).

LC–MS conditions

The nano LC-ESI/MS

Optimization of HPLC conditions for the separation of CA

Optimized chromatographic conditions are achieved after several trials with acetonitrile, water and formic acid in various proportions. It is found that the presence of formic acid in the mobile phase has a significant effect on the retention behavior of CA in Honeysuckle. Once 1% formic acid is applied instead of pure water, the pH is switched to 4.0, and then the tR (retention time) of CA on mobile phase is prolonged, avoiding eluting immediately when using water. In this HPLC system, an

Concluding remarks

An optimized MAE procedure has been developed and manifested more effective than other conventional techniques. For the first time, we report the extraction of CA from Honeysuckle using MAE and quantification of this compound by nano-LC-ESI/MS. The MAE method followed by nano-LC-ESI/MS determination, which takes comprehensive effect such as rapid analysis, reduced sample, time and solvent consumption, is a simple, rapid, and reliable method for the quality and quantify assessment of CA in

Acknowledgment

This paper was supported by 973 Program, 2007CB914100/3, 863 project: 2006AA02A308 and Shanghai Leading Academic Discipline Project, B109.

References (26)

  • W.C. Chang et al.

    Prostaglandins Leukot. Essent. Fatty Acids

    (1992)
  • X.G. Chen et al.

    J. Pharm. Biomed. Anal.

    (2006)
  • H.B. Xiao et al.

    J. Chromatogr. A

    (2004)
  • M. Gfrerer et al.

    Anal. Chim. Acta

    (2005)
  • E. Cortazar et al.

    Anal. Chim. Acta

    (2005)
  • B. Zhang et al.

    Sep. Purif. Technol.

    (2008)
  • X.J. Pan et al.

    Biochem. Eng. J.

    (2002)
  • N. Li et al.

    J. Chromatogr. A

    (2006)
  • C.H. Deng et al.

    J. Chromatogr. A

    (2006)
  • J.P.C. Vissers et al.

    J. Chromatogr. A

    (1997)
  • G. D’Orazio et al.

    J. Chromatogr. A

    (2005)
  • S.H. Hou et al.

    Talanta

    (2008)
  • R. Guo et al.

    Talanta

    (2008)
  • Cited by (64)

    • Enhanced extraction performance of iridoids, phenolic acids from Eucommia ulmoides leaves by tailor-made ternary deep eutectic solvent

      2021, Microchemical Journal
      Citation Excerpt :

      Isoquercetin, rutin and chlorogenic acid were seen as a class of compounds to experiment in this paper (Fig. S1). Despite of some researchers have investigated extract ingredients on technology of bioactive compounds in EUL, such as the extraction of individual components of chlorogenic acid from EUL [17,18], traditional methods to extract natural products from EUL will consume a large amount of volatile and toxic organic solvents, causing serious environmental pollution and threatening human beings [19]. With the development of studies, researchers also have used green solvents such as water and low-concentration ethanol to extract the active ingredients from EUL, while these methods require tough experimental conditions and are time-consuming [4,20].

    • Nanogold modified glassy carbon sensor for the quantification of phytoestrogenchlorogenic acid

      2020, Surfaces and Interfaces
      Citation Excerpt :

      According to several studies, CGA have been known to possess strong antioxidant activity and have beneficial health effects on human health such as it is an inhibitor of the tumor promoting activity of phorbol ester, it has potential role in regulating the glucose into the bloodstream in type-2-diabetes and appears to help people lose weight (metabolic disorder), phychoactive responses, liver function, Neurological conditions, Cancer, Cardiovascular diseases, Bacterial infections, oxidative and inflammatory diseases [11–15]. Techniques based on spectroscopy and chromatography such as potentiometric studies have acknowledged a great interest in researchers with novel study [16–33] Techniques traditionally used for the quantification of antioxidant properties of chlorogenic acid in biological samples and in natural samples includes high performance liquid chromatography (HPLC) [34], liquid chromatography mass spectrometry(LC-MS) [35], electrospray ionization mass spectrometry (EIS-MS) [36], capillary electrophoresis [37], NMR spectrometry [38] and ultra high performance liquid chromatography mass spectrometry (UHPLC-MS) [39]. However, these reported spectroscopic and chromatographic techniques offer high sensitivity but these methods have disadvantage that they are usually time consuming, require quite expensive equipment and pre-treatment steps prior to estimation of an analyte [40,41].

    View all citing articles on Scopus
    View full text