What Is Blueberry Extract Anthocyanin?

Blueberries (Vaccinium spp.) are in the genus Vaccinium of the family Ericaceae. The fruit is blue, nearly round, and 0.5–2.5 g in weight per fruit, with the largest being 5 g. Blueberries are one of the five healthy fruits recommended by the Food and Agriculture Organization of the United Nations. The earliest country to cultivate blueberries was the United States, and China began to import and cultivate them in 1981. Blueberries are rich in nutrients.

In addition to conventional nutrients, they also contain vitamin E, vitamin A, superoxide dismutase (SOD), anthocyanins and a large number of trace elements, such as calcium, phosphorus, magnesium, zinc, iron, germanium, copper, etc. 2. Blueberries are rich in anthocyanins is rich. Some literature reports that the anthocyanin content of blueberries is 387-487 mg/100 g, while that of blackberries is 245 mg/100 g, that of raspberries is 116 mg/100 g, and that of strawberries is 35 mg/100 g³. Anthocyanins have a variety of physiological functions. Extracting anthocyanins from blueberries and using them as dietary supplements not only makes full use of their physiological functions, but also solves the problem of blueberries' poor shelf life and perishability, and increases their commercial value. Therefore, in recent years, blueberry anthocyanins have received widespread attention.

1 Structure and properties of anthocyanins

1.1 Chemical structure

Anthocyanins are a type of flavonoid in the phenolic compound class. Their basic structure consists of two benzene rings connected by a -3 carbon unit (C6-C3-C6). The basic parent nucleus of anthocyanins is 2-phenylbenzopyran, the chromophore, as shown in Figure 1.4 Anthocyanins are usually glycosidically bound to one or more glucose, rhamnose, galactose, arabinose, etc. to form anthocyanins.

1.2 Types

More than 20 anthocyanins and over 250 anthocyanidins are known. There are six anthocyanins that are mainly found in the edible parts of plants: cyanidin, pelargonidin, peonidin, delphinidin, pelunidin and malvidin. (Malvidin), accounting for about 50%, 12%, 12%, 12%, 7% and 7% of the anthocyanin content in nature, respectively. Their chemical structures are shown in Table 1. 5. The main anthocyanins in blueberries are delphinidin, pelargonidin, peonidin, cyanidin and malvidin.

1.3 Chemical properties

Anthocyanin molecules contain acidic and basic groups and are soluble in water and alcohol compounds such as ethanol. They have strong absorption in both the ultraviolet and visible light regions, with the maximum absorption wavelengths near 280 nm and in the range of 500–550 nm. Anthocyanins appear in different colors at different pH values: red at pH<7, purple at pH 7–8, and blue at pH>11.

2. The physiological functions of blueberry anthocyanins

2.1 Antioxidant

A large number of studies have used chemical and cellular antioxidant methods to evaluate the antioxidant activity of blueberry anthocyanins, and the results show that blueberry anthocyanins have strong antioxidant activity. Wang Jian et al. evaluated the antioxidant capacity of blueberry anthocyanins through hydroxyl radical, DPPH radical, H₂O₂, superoxide anion radical and Fe³+ scavenging rate tests, and the results showed that blueberry anthocyanins have very high activity. Petko Denev et al. 8 tested the antioxidant properties of anthocyanin extracts from five types of berries: American cranberries, elderberries, redcurrants, blackberries and blueberries. The extracts were tested for their ability to absorb oxygen radicals (ORAC),hydroxyl radical scavenging capacity (HORAC), total hydroperoxyl radical-trapping antioxidant parameter (TRAP), NO scavenging and lipid peroxidation inhibition, it was found that blueberry extract had the strongest HORAC and NO scavenging abilities.

Spela Moze Bornsek et al. 9 conducted intracellular antioxidant assays (CAA) in different cell lines such as human colon cancer (Caco-2), liver cancer (HepG2), human endothelial cells (EA.hy926), and rat vascular smooth muscle cells (A7r5), and the results showed that anthocyanins have intracellular antioxidant properties. Sun Liqiong et al. studied the antioxidant activity of anthocyanin monomers in 10 of the 9 Chinese-introduced blueberry varieties. The results showed that delphinidin and cyanidin-3-glucoside are the main active ingredients in blueberries.

2.2 Protects eyesight

Blueberry anthocyanins are an important element in protecting eyesight. They can protect the capillaries in the eyes, thereby promoting blood circulation; and they can accelerate the regeneration of rhodopsin, which is indispensable for good eyesight. Blueberry anthocyanins can significantly improve blurred vision, dry eyes, visual acuity and overall efficiency, thereby relieving visual fatigue. Children with mild myopia can effectively control and alleviate the development of myopia by supplementing 500 mg of blueberry anthocyanins daily[3].

2.3 Lower blood sugar

Male obese rats were given 200 mg/(kg·d) blueberry anthocyanins by gavage. After 4 weeks, fasting blood was collected to measure blood glucose. The results showed that compared with the control group, blueberry anthocyanin treatment significantly reduced the fasting blood glucose level of rats, and blueberry anthocyanins could improve the insulin sensitivity. 4. Grace MH et al. 15 gave mice blueberry anthocyanins, which lowered their blood glucose levels by 33% to 51%, while the blood glucose levels of mice given metformin were reduced by 27%, indicating that blueberry anthocyanins can effectively alleviate the symptoms of hyperglycemia in mice.

2.4 Anti-proliferation

Bunea A et al. [0 showed that blueberry anthocyanins have anti-proliferative and pro-apoptotic effects on the mouse melanoma cell line B16-F10. The four concentrations of blueberry anthocyanins, 50, 100, 150, and 200 μg/mL, were selected and applied to human glomerular mesangial cells for 24 hours. The inhibition rates of the cells were 40.98%, 60.39%, 59.60%, and 64.66%, respectively, indicating that blueberry anthocyanins inhibit cell proliferation in a dose-dependent manner and induce apoptosis.

2.5 Other physiological functions

In recent years, scholars have discovered that in addition to the above physiological functions, blueberry anthocyanins also have other functions, such as in vitro inhibition of hepatitis B virus, anti-liver damage, anti-radiation damage, analgesic and anti-inflammatory, and anxiety relief 18-23].

3 Factors affecting the stability of blueberry anthocyanins

The stability of blueberry anthocyanins is poor. Processing conditions such as pH, temperature, light, metal ions and additives can affect their stability and hinder their application. Blueberry anthocyanins are stable under acidic conditions. High temperatures and light can accelerate their degradation. The effect of metal ions and sugars on their stability is related to their concentration. Therefore, alkaline and high-temperature environments should be avoided as much as possible during processing, and they should be stored under low-temperature, light-protected conditions.

3.1 pH

pH has a significant effect on the stability of blueberry anthocyanins. When pH<3, blueberry anthocyanins are relatively stable; when pH>3, the anthocyanin structure is unstable and easily degraded. 4; The molecular structure of anthocyanins at different pH values will change, which will lead to a change in color. 2.

3.2 Temperature

The stability of blueberry anthocyanins decreases significantly at high temperatures, and the degradation rate of blueberry anthocyanins accelerates with increasing temperature. Liu Junbo et al. 20 showed that when below 60 °C, the stability of blueberry anthocyanins is less affected. Wang Bochu et al. 27 showed that blueberry anthocyanins are sensitive to high temperatures (≥ 80 °C).

3.3 Light

Blueberry anthocyanins are unstable in light. Studies have shown that anthocyanins are unstable and easily degraded in natural light, and are more stable in dark conditions. Therefore, blueberry anthocyanins should be stored in the dark.

3.4 Metal ions

Metal ions are often introduced during processing, and these metal ions can affect the function of anthocyanins. Therefore, it is important to study the effect of metal ions on the stability of anthocyanins. The results showed that Fe³+, Fe²+, and Cu²+ affected the stability of anthocyanins. Zn²+ ions with a concentration greater than 1.2 mmol/L, Cu₂+ with a concentration greater than 12 mmol/L, and Ca²+ with a concentration greater than 12 mmol/L were beneficial to the stability of blueberry anthocyanins. Na+ had no significant effect on the stability of blueberry anthocyanins

3.5 Oxidants and reducing agents

Oxidants and reducing agents are used in the processing. Blueberry anthocyanins are sensitive to oxidants (H₂O₂, 0.5%~2.0%) and reducing agents (Na₂SO₃, 0.005~0.040 mol/L). Sun Qianyi Sun et al. 24 found that as the concentration of the oxidant and reducing agent increased, the preservation rate of blueberry anthocyanins decreased rapidly. At a concentration of 2% H₂O₂ and Na₂SO₃, the preservation rates of blueberry anthocyanins were low, 34% and 35%, respectively.

3.6 Sugars

Sugars are widely used in food processing. Commonly used sugars include glucose, sucrose and fructose. The effect of sugars on the stability of blueberry anthocyanins is related to the concentration of the sugars used. The results of a study by Sun Qianyi et al. 24 showed that glucose, sucrose and fructose at concentrations of more than 20% were stable to blueberry anthocyanins. A study by Yan Hongguang et al. 25 showed that sucrose and glucose at concentrations higher than 10% had a slight protective effect on anthocyanins, while low concentrations of sucrose and fructose can reduce the stability of blueberry anthocyanins. Liu Junbo et al. 20 showed that sucrose concentrations below 50g/L (5%) had no significant effect on blueberry anthocyanins, while high-quality sucrose concentrations could improve stability. These results indicate that high concentrations of sugars are beneficial to the stability of blueberry anthocyanins.

3.7 Other food additives

Food additives can prolong the shelf life of food, while also improving the sensory properties of food to meet people's sensory needs, and are widely used in the food processing industry. Research results show that VC can damage the structure of anthocyanins and degrade anthocyanins, while sodium benzoate, 0.15% citric acid and malic acid can promote the stability of blueberry anthocyanins.

3.8 Other factors

In addition to the above factors that affect the stability of blueberry anthocyanins during food processing, some processing conditions can also affect the stability of blueberry anthocyanins. For example, Nadiarid Jiménez et al. 28 determined the degradation rate of anthocyanins in blackberry juice samples at three different water activities. The results showed that as the water activity decreased from 0.99 to 0.34, the degradation rate constant of anthocyanins increased from 0.9×10-3/s to 3.5×10-3/s, which indicates that a decrease in water activity has a negative impact on the stability of anthocyanins at high temperatures.

4 Extraction and purification of blueberry anthocyanins

4.1 Extraction methods

At present, the main methods for extracting blueberry anthocyanins include organic solvent extraction, ultrasonic-assisted extraction, microwave-assisted extraction and enzymatic methods. Some methods combine several methods, such as ultrasonic-enzymatic extraction and ultrasonic-microwave extraction.

4.1.1 Solvent extraction method

Solvent extraction is one of the most commonly used methods for extracting anthocyanins. As anthocyanins are soluble in alcohol solutions such as ethanol and methanol, ethanol or methanol is usually used as the extraction agent. Anthocyanins are stable under acidic conditions, so blueberry anthocyanins are usually extracted under acidic conditions.

A methanol solution is a commonly used solvent for extracting blueberry anthocyanins. A small amount of hydrochloric acid or acetic acid solution can be added to improve the extraction efficiency. The best process conditions can be obtained by adjusting the liquid-to-material ratio, extraction temperature and extraction time. 2930 However, methanol solutions are toxic and not environmentally friendly, so ethanol solutions are often used as extraction solvents. [3]

4.1.2 Ultrasonic extraction method

Ultrasound causes the cell walls of the blueberries and the entire blueberry fruit to break in an instant, shortening the crushing time. At the same time, the vibration generated by ultrasound enhances the release, diffusion and dissolution of intracellular substances, thereby significantly improving the extraction efficiency. Wu Jinming et al. 3 used response surface methodology to analyze and optimize the ultrasonic extraction process of blueberry anthocyanins, and concluded that the optimal extraction conditions for blueberry anthocyanins were ultrasonic power 730 W, liquid-to-material ratio 1:18, extraction time 40 min, and extraction temperature 55 °C. Under these conditions, the anthocyanin extraction rate was 5.79%.

4.1.3 Enzymatic method

Enzymes can break down plant tissues, thereby accelerating the release of active substances in plant cells and thus improving the extraction rate. Zhao Erfeng et al.133 used cellulase to extract anthocyanins from blueberry pomace, and the anthocyanin extraction rate of the resulting blueberry pomace was 4.12%.

4.1.4 Other methods

Boyaqiu A compared the effects of solvent extraction and microwave-assisted extraction on the extraction rate of anthocyanins. The results showed that microwave-assisted extraction had a better extraction effect. Michael Schwarz et al. 3 confirmed the potential of high-speed countercurrent chromatography for large-scale separation of anthocyanins. High-speed countercurrent chromatography has several advantages: high sample load, low separation solvent cost, and does not require expensive solid phase extraction columns. The operating conditions are also gentle. Wang Erlei et al.3 used column chromatography combined with semi-preparative high performance liquid chromatography to successfully isolate a high-purity anthocyanin mixture and anthocyanin monomers from wild blueberries. An effective elution system for separating high-purity anthocyanin compounds was established: First, an Amberlite XAD-7HP column was filled with an aqueous ethanol solution containing 0.01% hydrochloric acid (ethanol: water = 35:65), and then a Sephadex LH-20 column was filled with an aqueous ethanol solution containing 0.01% hydrochloric acid (ethanol: water = 35:65).

4.2 Purification method

The crude extract of blueberry anthocyanins contains many impurities, such as proteins, sugars, pectin and other substances. In order to improve the quality and extend the shelf life of the product, it is necessary to purify the crude extract of blueberry anthocyanins.

4.2.1 Resin method

The adsorption separation technology of macroporous resin is a new extraction and purification process that uses a special adsorbent to selectively adsorb the active ingredients and remove the inactive ingredients. This method has the advantages of simple equipment, convenient operation, energy saving, low cost, high product purity, and no moisture absorption. Currently, the resin method is the main purification method for blueberry anthocyanins. Timothy J. Buran and others37 found that Amberlite FPX66 resin is the most suitable resin for recovering anthocyanins and flavonoids from blueberry water extracts.

4.2.2 Solid phase extraction method

Petko Denev et al. 8 used solid phase extraction (SPE) to purify anthocyanins from five berries (American chokeberry, elderberry, red currant, blackberry and blueberry). The results showed that solid phase extraction (SPE) using the absorbent Amberlite XAD7 is a fast, simple and reliable method for purifying anthocyanins from complex extracts containing impurities such as sugars, acids, proteins and pectin.

5 Development prospects

Blueberries are a good source of anthocyanins due to their high anthocyanin content and good functionality. They have been widely used in the food, medical and cosmetic industries. However, blueberry anthocyanins are unstable and are lost during the production process. The problem of how to retain anthocyanins and their physiological activity to the greatest extent possible needs to be urgently solved. Anthocyanins also have poor stability in the human gastrointestinal tract and low bioavailability. However, the biological properties of anthocyanins are mainly determined by their bioavailability, so improving the bioavailability of anthocyanins is also a key area of future research.

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