If there is no normal macular area, people's main visual function will be gradually damaged, and even there is a danger of blindness. In the center of macular region, the incident light is the strongest and the most reactive oxygen species are produced. A large number of epidemiological studies also show that zeaxanthin can specifically absorb the blue light that is most harmful to the retina, thus protecting the cone cells in the fovea. Many studies have shown that increasing the intake of zeaxanthin in the short term can increase the macular pigment, thus enhancing the ability of macular area to resist harmful substances and light damage, and preventing and slowing down age-related macular degeneration.
In addition, zeaxanthin itself has high nutritional value, which can be converted into bioactive vitamin A in human liver after eating, and has special effects on promoting human growth and development, protecting vision and epithelial cells, improving disease resistance and prolonging life. Zeaxanthin mainly exists in the epidermis of yellow corn. Corn processing by-products that can be used to produce zeaxanthin include corn flour, DDGS and corn bran. Extraction techniques include organic solvent extraction, ultrasonic extraction, microwave-assisted extraction, surfactant extraction, enzyme extraction, supercritical fluid extraction and membrane-assisted separation extraction.
Organic solvent extraction method
The method takes single solvent or mixed organic solvent such as petroleum ether, ethanol and acetone as extractant, mixes the raw materials to be treated with the extractant, slowly stirs and extracts for several hours at room temperature, separates the mixed oil from the leached material, and recovers the solvent in the mixed oil to obtain a carotenoid mixture containing zeaxanthin, cryptoxanthin and lutein. The main characteristics of organic solvent separation and extraction method are relatively simple extraction process and high extraction rate. The filter residue obtained by filtration in the process can be leached twice, and the solvent obtained after distillation can be recovered and extracted. This method especially needs to grasp the extraction time. The extraction time is too short, the extraction is insufficient, and the extraction time is too long, which is easy to precipitate other impurities and affect the purity.
Enzymatic extraction
Carotenoids and protein in plants generally exist in a combined state. The crude zeaxanthin obtained by traditional direct extraction and concentration contains a certain amount of gliadin, which is not conducive to the final purification of zeaxanthin. Enzymatic extraction of zeaxanthin is to hydrolyze part of protein with protease and break the network structure of protein, which can not only improve the extraction rate of zeaxanthin, but also obtain higher purity zeaxanthin. When extracting zeaxanthin by enzymatic method, it is necessary to control the substrate concentration, enzyme concentration, pH, hydrolysis time and hydrolysis temperature during the hydrolysis of corn gluten meal.
Microwave-assisted extraction
Microwave-assisted extraction technology is a new extraction technology developed on the basis of traditional solvent extraction principle. Microwave extraction can strengthen the extraction process, reduce production time, energy, solvent consumption and waste generation, improve yield, reduce operating costs, and meet the requirements of environmental protection. This is a new technology with good development prospects. Microwave-assisted extraction of maize yellow pigment has the advantages of short time, high extraction rate, low solvent consumption, high recovery rate, energy saving and environmental pollution reduction.
Surfactant extraction method
Surfactant extraction of zeaxanthin is also a method based on organic solvent extraction and microwave-assisted extraction technology. With the help of surface method, the pollution of organic solvents to pigment products is reduced, which has the advantages of fast speed and high extraction rate, and provides a new way for the development and utilization of corn yellow pigment.
Ultrasonic extraction
This method can improve the yield of zeaxanthin based on direct extraction and ultrasonic wave. In the process of extracting ultrasonic radiation, the acoustic energy of ultrasonic field produces ultrasonic cavitation, which greatly accelerates the internal diffusion speed. At the same time, the surface of solid particles is stripped, eroded and crushed to produce new active surface, which increases the specific surface area of mass transfer and thus improves the extraction speed. Ultrasonic extraction of zeaxanthin shortens the extraction time, and the operation is simple, and the quality of the obtained product is stable, but the protein content of the extract is high, so it is necessary to further purify and separate the product to obtain zeaxanthin.
Supercritical fluid extraction
Supercritical fluid extraction (SFE) device is a special solid-liquid or liquid-liquid separation device, and the commonly used extraction solvents are CO2 and propane. Supercritical fluid extraction of zeaxanthin does not introduce any chemical synthetic substances, and the operating temperature is low, the pressure is not too high, and the zeaxanthin is not easy to degrade, which is conducive to maintaining the naturalness of zeaxanthin. However, due to the high requirements for equipment, large-scale industrial production cannot be formed and the output rate is low. Membrane-assisted separation and extraction technology
Membrane separation, as a newly developed high-tech separation technology, uses a semi-permeable membrane as a barrier layer to separate and purify different components in the mixture under the action of energy, concentration or chemical potential difference with the help of membrane selective permeation. The whole process device is relatively simple, and has the advantages of convenient operation, compact structure, low maintenance cost, convenience and easy automatic control. Zeaxanthin belongs to isoprene and often coexists with cryptoxanthin, β -carotene and lutein. To form a carotenoid mixture. The commonly used separation and purification methods are thin layer chromatography and high performance liquid chromatography.
Thin chromatographic separation method
Thin-layer chromatography is one of chromatographic analysis methods, which has the characteristics of simple instrument and operation, fast development time and high detection sensitivity. It is not only suitable for the separation and identification of trace components, but also suitable for the preparation of a small amount of pure substances. Thin layer chromatography can be used to separate, refine and identify various types of compounds, such as inorganic substances, organic substances, small or large molecular compounds, hydrophilic substances or lipophilic substances. Zeaxanthin belongs to isoprene pigment and is mainly composed of carotenoids such as zeaxanthin and lutein. Therefore, zeaxanthin was separated by adsorption and thin layer chromatography, and the separated components were qualitatively identified by infrared and ultraviolet spectra.
high performance liquid chromatography
The quantitative analysis of lutein and zeaxanthin in food generally adopts electron (ultraviolet-visible) absorption spectrometry. Because they exist in many food raw materials, their separation is the basis of quantitative analysis. During 1994, Sander et al. successfully separated carotenoids and their geometric isomers on C30 stationary phase for the first time on high performance liquid chromatography. Since then, C30 column has been increasingly used for the separation and detection of carotenoids. Researchers applied C30 column to high-pressure liquid chromatography (HPLC) equipped with diode array detector (PDA), so that all-trans lutein and zeaxanthin in food were well separated. Zeaxanthin can be identified according to its chromatographic behavior and spectral characteristics.
Preparation and synthesis method Biosynthesis method
Zeaxanthin is a secondary metabolite of plants. Zeaxanthin was synthesized by regulating isoprene pathway by recombinant gene technology, and high-yield plants or strains were constructed. Therefore, there are two ways to biosynthesize zeaxanthin: cultivating a large number of fungi that can synthesize zeaxanthin and constructing plants or strains with high yield of zeaxanthin through genetic technology.
Foreign researchers cloned the gene cluster of Agropyron bicolor into Escherichia coli and expressed it in Saccharomyces cerevisiae cells after transformation. After deleting some redundant parts, the GGPP enzyme activity expressed by the recombinant gene increased from 6.35 to 23. 4 nanomoles per minute. The initial coding GTG of lycopene cyclase was replaced by ATG, and the gene encoding phytofluene synthase was fused with the phosphoglycerate kinase promoter of Saccharomyces cerevisiae, and the yeast was transformed with the integrated vector, which could produce 5% zeaxanthin. In 2000, the researchers connected two genes, PSY and LycB, to the endosperm-specific glutenin promoter, and at the same time connected the bacterial phytoene degrading enzyme gene crtI to the cauliflower spot virus 35S promoter, then constructed the expression vector and transferred it to Japanese rice varieties. As a result, zeaxanthin was detected in rice endosperm. Isomerization is the preparation of zeaxanthin isomers by chemical methods. Because zeaxanthin and lutein are isomers, zeaxanthin can be produced by converting lutein in the presence of polyol and alkali. Isomerization of lutein to zeaxanthin is faster, and the whole process can be carried out under normal pressure, which is more suitable for industrial production.
Kar rer and Jucker reported that lutein can be converted into zeaxanthin in the presence of sodium ethoxide and benzene. Andrews also reported that lutein can be transformed into zeaxanthin in the presence of methanol, potassium methoxide and dimethyl sulfoxide in nitrogen. The U.S. patent reports a method of isomerizing lutein in an aqueous system to produce zeaxanthin without a catalyst. In this method, lutein reacts with strong alkali aqueous solution for a long time to obtain zeaxanthin. The determination methods of zeaxanthin mainly include spectrophotometry and high performance liquid chromatography. When using the spectrophotometer method, take zeaxanthin extract, take the extracted organic solvent as the reference, and directly measure the absorption value in the UV-Vis spectrophotometer, the wavelength is generally 445 nm, and judge the zeaxanthin content according to the absorption value. The sensitivity detection limit of spectrophotometer method can reach 105~ 106, and the sample volume is 100? L order of magnitude. Because of its simple operation and low cost, this method can be used to screen corn materials rich in zeaxanthin.
High performance liquid chromatography (HPLC) can determine various components in the extract samples, including various carotenoids, such as lutein and zeaxanthin. This method has high sensitivity. For example, the sensitivity of fluorescence detector can reach 10~ 1 1 g and the detection limit is 109. What is the sample size? L order of magnitude, quantitative detection is not affected by impurities in the extract. However, this method requires high instruments, requires specialized personnel to operate, and is expensive, so it is difficult to be applied in ordinary laboratories, and can be used as a later quantitative determination of maize rich in zeaxanthin.