CROSS-REFERENCE TO RELATED APPLICATIONS

This application claims priority to Chinese Patent Application No. 201911021945.0, filed on Oct. 25, 2019, the entire contents of which are incorporated herein by reference.

TECHNICAL FIELD

The present invention relates to the field of detection technique, and in particular to a reference fingerprint of virgin olive oil, and an establishment method and an application thereof.

BACKGROUND

Olive oil is a type of oil obtained from fresh olives (not including oils obtained by solvent extraction or re-esterification process) and undoped with other types of oils and fats. Virgin olive oil is of the highest quality and nutritive value, which is rich in such natural active substances as unsaturated fatty acids, squalene, polyphenol, and vitamins, and plays roles in antioxidation, cholesterol regulation, cancer prevention, and cosmetology, with far higher price compared with other edible vegetable oils. With unique physicochemical properties and superb nutritional and healthcare effects, olive oil is favored by people. In view of this, some merchants are prompted by economic interests, resulting in always serious problems with adulterate olive oil. So far, there are two most pervasive types of problems with adulteration of olive oil: one is that cheap vegetable oil is mixed with expensive olive oil; the other is that low-grade virgin olive oil or pomace olive oil is added into extra virgin olive oil.

Because of difficulty in adulteration identification of olive oil, related testing and trade standards are not far enough to satisfy practical requirements. Conventional adulteration identification and detection method of virgin olive oil is to determine the quality or identity of the olive oil by means of detection of quality and characteristic indexes (acid value, peroxide value, residual solvent, fatty acid composition, unsaponifiable matters, and ultraviolet absorbance). In addition, one of the most commonly used standard method for identifying adulteration of olive oil internationally is sterol analysis. However, this method has problems with complex sample pretreatment process, many interference factors, poor reproducibility, and long detection cycle. Olive oil identification methods reported include near-infrared spectroscopy, determination of the wax content, nuclear magnetic resonance (NMR) spectroscopy, and GC-MS method. All of these methods play active roles to some extent, but the complexity of adulteration of olive oil and different indexes of other adulterated vegetable oils challenge the foregoing analytical method and often bring a lot of confusion to identification. Therefore, it is necessary to develop a relatively stable, practical, and highly accurate adulteration identification method with reference to the fingerprint of virgin olive oil, in order to fight against such undesirable phenomena as shoddy and adulterate olive oil.

A standard fingerprint is a spectrum representing common characteristics of fingerprints of a batch of samples of the same type. Research on fatty acid fingerprints is the most widely and deeply studied so far. However, some studies have shown that some vegetable oils have a similar fatty acid composition. Therefore, there is some risk associated with identification and characterization of these oils and fats with similar fatty acid composition by means of fatty acid fingerprints; also, using fatty acid fingerprints to identify adulterate oils merely has the possibility of misjudgment because the type and extent of adulteration are also large.

SUMMARY

In view of this, the objective of the present invention is to provide a reference fingerprint of virgin olive oil, and an establishment method and an application thereof. The reference fingerprint of virgin olive oil provided by the present invention can evaluate the quality of virgin olive oil rapidly and accurately.

To achieve the above purpose, the present invention provides the following technical solution.

The present invention provides an establishment method of a reference fingerprint of virgin olive oil, including the following steps:

mixing 10-30 batches of extra virgin olive oil with a solvent for extraction, to obtain serial extra virgin olive oil samples;

conducting ultra-performance liquid chromatography-fluorescence detection (UPLC-FLD) on the extra virgin olive oil samples, to obtain the chromatographic information of the serial extra virgin olive oil samples;

conducting similarity calculation on the chromatographic information of the serial extra virgin olive oil samples, and obtaining the reference fingerprint of the virgin olive oil.

Preferably, the solvent is methanol, acetonitrile, or ethanol.

Preferably, the consumption ratio of the extra virgin olive oil to the solvent is 1.0-5.0 g:5-20 mL.

Preferably, conditions of the UPLC-FLD include:

column: RP-C18column or SVEA C18column;

flow rate: 0.1-0.4 mL/min;

excitation wavelength: 300 nm;

emission wavelength: 350 nm;

column temperature: 25-40° C.;

injection volume: 0.1-3.0 μL;

mobile phase: A is 0.0-0.3% (v/v) formic acid-methanol or 0.0-0.3% (v/v) formic acid-acetonitrile; B is 0.05-0.3% (v/v) formic acid-water;

the gradient elution program is:

0-3 min: 10% A, 90% B; and

3-45 min: 55% A, 45% B.

Preferably, the software for similarity calculation is Similarity Evaluation System for Chromatographic Fingerprint of TCM (Version A) published by the Chinese Pharmacopoeia Commission.

Preferably, the area of a fingerprint peak in the reference fingerprint of virgin olive oil is greater than 5%.

The present invention further provides a reference fingerprint of virgin olive oil obtained by the above establishment method.

The present invention further provides an application of the above reference fingerprint of virgin olive oil in the field of detection of quality of virgin olive oil.

Preferably, the application includes the following steps:

mixing the virgin olive oil with a solvent for extraction, to obtain a virgin olive oil solution;

conducting ultra-performance liquid chromatography-fluorescence detection (UPLC-FLD) on the virgin olive oil solution, to obtain the chromatographic information of the virgin olive oil solution;

with reference to the reference fingerprint of virgin olive oil, conducting similarity calculation on the chromatographic information of the virgin olive oil solution, to achieve the purpose of evaluating the quality of virgin olive oil.

Preferably, the application further includes:

dissolving coumarin, hydroxytyrosol, vanillic acid, and salicylic acid standards in a solvent, to obtain a mixed standard solution of coumarin, hydroxytyrosol, vanillic acid, and salicylic acid;

conducting UPLC-FLD on the mixed standard solution, to obtain the chromatographic information of the mixed standard solution;

with reference to the chromatographic information of the mixed standard solution, assigning the chromatographic information of the virgin olive oil solution.

The present invention provides an establishment method of reference fingerprint of virgin olive oil, including the following steps: mixing 10-30 batches of extra virgin olive oil with a solvent for extraction, to obtain serial extra virgin olive oil samples; conducting ultra-performance liquid chromatography-fluorescence detection (UPLC-FLD) on the extra virgin olive oil samples, to obtain the chromatographic information of the serial extra virgin olive oil samples; conducting similarity calculation on the chromatographic information of the serial extra virgin olive oil samples, and obtaining the reference fingerprint of the virgin olive oil. In the present invention, serial extra virgin olive oil samples are analyzed using UPLC-FLD, and the chromatographic information of various components in the extra virgin olive oil is obtained, breaking through the limitation of establishment of fingerprints by means of conventional GC-MS method and based on the fatty acid composition of edible oil, and further solving the problem that traditional fingerprints only use a single component for determination and thus result in false positivity. Meanwhile, similarity calculation is conducted on the serial extra virgin olive oil, establishing a reference fingerprint of virgin olive oil based on common peaks, reinforcing the polymorphism of reference fingerprint information, reflecting its differences from edible oil from the perspectives of the substance composition, distribution, and content of virgin olive oil, helping improve accuracy of identification, and enabling rapid, convenient and accurate identification of the quality of virgin olive oil. It possesses high application value in routine quality analysis of virgin olive oil and inspections for market supervision and industrial production.

The present invention further provides an application of the reference fingerprint of virgin olive oil provided in the foregoing technical solution in the field of detection of quality of virgin olive oil. The reference fingerprint of virgin olive oil established by the present invention can evaluate the quality of virgin olive oil more sensitively and accurately due to a number of reference peaks.

