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Determination of Crude Polysaccharides by Ultraviolet Spectrophotometry
Keywords: UV spectrophotometry; Lycium barbarum polysaccharide; Ganoderma lucidum polysaccharide; Aesthetic analysis instrument
The content of crude polysaccharides in food was measured using the sulfuric acid-phenol colorimetric method, with glucose as a standard and dextran as an alternative. The results showed that the former method yielded slightly higher values, approximately 4.8% higher. This technique is simple, fast, accurate, and highly reproducible, making it suitable for determining various types of crude polysaccharides.
A polysaccharide is defined as a carbohydrate composed of ten or more monosaccharides linked by glycosidic bonds. These are typically natural polymers and can be categorized into active polysaccharides and dietary fiber. Active polysaccharides, such as those found in fungi, plants, and chitosan, exhibit a wide range of physiological functions, including immune regulation, anti-cancer effects, anti-aging properties, and lipid-lowering capabilities. Despite their importance, there are no standardized national methods for polysaccharide determination, and most literature reports rely on the phenol-sulfuric acid or anthrone-sulfuric acid methods.
Most standard curves are prepared using glucose as the reference rather than dextran due to its availability and cost. However, research has shown that polysaccharides from mushrooms, Flammulina, Yunzhi, Dongcao, Xia Cao, Ganoderma lucidum, Poria, and Polyporus have significant biological activities, such as immune enhancement, tumor suppression, sedative effects, and anti-inflammatory properties.
This study compared the use of glucose and dextran as standards for determining the polysaccharide content in foods. It involved two types of polysaccharides: Lycium barbarum polysaccharide and Ganoderma lucidum polysaccharide.
**Materials and Methods**
**1.1 Test Materials**
- Lycium barbarum polysaccharide (40–60% content, tested by national standards)
- Ganoderma lucidum polysaccharide (40–60% content, tested by national standards)
- Dextran (molecular weight 500,000)
**1.2 Reagents**
- Copper stock solution
- Copper reagent solution
- Detergent
- Ethanol solution (80%)
- Sodium hydroxide solution (10%)
- Sulfuric acid solution
- Phenol solution (5%)
- Glucose/dextran standard stock solution
- Glucose/dextran standard working solution
**1.3 Instruments**
- UV spectrophotometer UV-1100
- UV-1200 centrifuge rotary mixer
**1.4 Experimental Method**
**1.4.1 Preparation of Glucose Standard Curve**
- Standard solutions were prepared with concentrations of 0, 0.02, 0.04, 0.06, 0.08, and 0.10 mg/ml.
- Absorbance was measured at 490 nm using a 1 cm cuvette.
- The regression equation was Y = 4.4122X – 0.0147, with a correlation coefficient r = 0.9996.
**1.4.2 Preparation of Dextran Standard Curve**
- Similar procedure was followed with dextran solutions.
- The regression equation was Y = 4.635X – 0.0145, with r = 0.9983.
**1.5 Sample Determination**
**1.5.1 Sample Extraction**
- 2.0 g of crude polysaccharide was dissolved in 100 ml water and heated for 2 hours.
- After cooling, the solution was filtered and used for further steps.
**1.5.2 Precipitation of Crude Polysaccharides**
- 1.5 ml or 4.0 ml of filtrate was mixed with 16 ml ethanol and centrifuged.
- The precipitate was washed and dissolved in 5 ml water.
**1.5.3 Precipitation of Dextran**
- 2 ml of the solution was treated with NaOH and copper reagent, then boiled and centrifuged.
- The precipitate was dissolved in sulfuric acid and diluted to 50 ml.
**1.5.4 Determination of Sample**
- 2 ml of sample solution was mixed with phenol and concentrated sulfuric acid.
- Absorbance was measured at 490 nm, and the concentration was calculated based on the standard curve.
**Stability Test**
- The absorbance was measured at intervals after color development, showing stable results over time.
**Recovery Rate**
- The recovery rate ranged between 93.15% and 99.7%, indicating good accuracy.
**Summary**
- Using glucose instead of dextran as a standard is more practical in China due to cost and availability.
- The method allows for selective precipitation of high molecular weight polysaccharides.
- No need for purification or decolorization makes this method efficient and reliable.
- Proper washing of the precipitate is essential to avoid interference from other sugars.
Keywords: ultraviolet spectrophotometry; alfalfa polysaccharide; ganoderma lucidum polysaccharide; aesthetic analyzer; UV-1100; UV-1200
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