Jun 09, 2025
To Perform Quantitative estimation of carbohydrates using Dinitro Salicylic Acid (DNS Method)
- Durvesh Smita. Burhade1,
- Paridhi Jain1,
- Lakshay Saini1,
- Yash Modi1
- 1Parul University
- Durvesh Smita. Burhade: Project Leader
- Durvesh Burhade Lab
Protocol Citation: Durvesh Smita. Burhade, Paridhi Jain, Lakshay Saini, Yash Modi 2025. To Perform Quantitative estimation of carbohydrates using Dinitro Salicylic Acid (DNS Method). protocols.io https://dx.doi.org/10.17504/protocols.io.3byl41o3olo5/v1
Manuscript citation:
Burhade, D., Jain, P., Saini, L., Mathur, V., & Modi, Y. (2025). Comprehensive Review of Fundamental Biochemical Techniques: Integration of Quantification, Separation, and Analytical Methods in Biomolecular Studies (1.2). Open Access 2025. https://doi.org/10.5281/zenodo.14872899
License: This is an open access protocol distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited
Protocol status: Working
We use this protocol and it's working
Created: June 07, 2025
Last Modified: June 09, 2025
Protocol Integer ID: 219741
Keywords: Quantitative Estimation, DNS Method
Funders Acknowledgements:
Yash Modi
Grant ID: ORP-BIOCHEMISTRY-2024/25
Abstract
This method is used for the measurement of total carbohydrates present in the given sample. Quantitative analysis of carbohydrates is important for a variety of reasons, like nutritional labelling, quality control, authenticity, caloric content, and energy evaluation. We use Dinitro salicylic acid,which is a low carcinogenic reagent and light sensitive and also used to bind functional groups of glucose in alkaline conditions. Here we are doing a quantitative estimation of sugar or carbohydrate. DNS is used only in alkaline conditions. We can also use this method to determine the concentration of an unknown sugar sample by preparing a set of solutions with known C6H12O6 concentration. Then, plot a standard curve and use it to derive the concentrations of unknown samples. There are some properties of reducing sugars that reduce so many reagents DNS is also one of them, so we used to perform quantitative estimation, which is used to determine the mass, percentage composition or concentration of a substance.
Image Attribution
Reduction of DNS (3,5-dinitrosalicylic acid) to 3 amino, 5-nitro salicylic acid.
Guidelines
The results demonstrate a clear inverse relationship between sugar concentration and absorbance, with higher sugar levels yielding stronger colourimetric responses. Tube 1’s extreme absorbance suggests potential reagent saturation, which may require sample dilution for accurate quantification in future assays. The data confirms the DNS method’s effectiveness in detecting a wide range of reducing sugar concentrations. The results clearly demonstrate that higher sugar concentrations lead to greater absorbance at 540 nm. The extreme value in Tube 1 may require dilution for accurate quantification, as absorbance values above 1.5 often exceed the linear detection range of spectrophotometers.
Materials
| Component | Description | |
| Sodium sulphate solution | Use to make DNS reagent | |
| Distilled water | Use to make DNS reagent | |
| 3,5-dinitrosalicylic acid | Use to make DNS reagent | |
| Sodium hydroxide | Use to make DNS reagent | |
| Sodium potassium titrate | Use to make DNS reagent |
Materials and components required for this experiment
| A | B | |
| Glassware | Description | |
| Test tube | Used to store a solution. | |
| Pipette | Used to measure the volume of a solution. | |
| Boiling water bath | To keep the solution at a constant temperature. |
Glassware Required for this experiment
Safety warnings
Wear a Lab Coat & Purple Nitrile Gloves while handling solutions
DNS Solution & Quantitative estimation using sugar as a Source
DNS Solution & Quantitative estimation using sugar as a Source
Take 7 clean, dry test tubes
Pipette out standard sugar solution in different test tubes and make up the volume of all test tubes to 1 mL with distilled water.
Add 2 mL DNS reagent to all the test tubes. Mix the plug in the test tube with cotton or marble and keep the test tube in a boiling water bath for 5 minutes.
Take the tubes and cool them to Room temperature
Add 7 ml of water after cooling the solution completely.
Read extinction at 540 nm against the blank.
Take Blank Water
All the tubes must be cooled to room temperature before reading since the absorbance is sensitive to temperature.
we use carbohydrates of different concentrations to measure absorbance of sugar
Observation Table
Results
Protocol references
To Learn more about this method, read this paper
Citation of the Article:
Burhade, D., Jain, P., Saini, L., Mathur, V., & Modi, Y. (2025). Comprehensive Review of Fundamental Biochemical Techniques: Integration of
Quantification, Separation, and Analytical Methods in Biomolecular Studies (1.2). Open Access 2025. https://doi.org/10.5281/zenodo.14872899