A Comprehensive Guide to the Methods to Assess Protein Quality and Quantity

January 11, 2026 •

4 min read

Accurate protein quantification is critical for ensuring proper nutritional labeling, with the Kjeldahl method historically serving as the industry standard. This comprehensive guide explores the primary laboratory and bioassay methods to assess protein quality and quantity, clarifying how food scientists evaluate this essential macronutrient.

Quick Summary

This article details the scientific techniques used for protein analysis, covering quantitative measures like the Kjeldahl and Dumas methods and qualitative assessments such as the DIAAS and PDCAAS scoring systems.

Key Points

Protein Quantity vs. Quality: Quantity measures total protein content (e.g., Kjeldahl, Dumas), while quality assesses its nutritional value based on amino acid profile and digestibility (e.g., DIAAS, PDCAAS).
Kjeldahl and Dumas: The Kjeldahl method is a traditional, robust but slow nitrogen-based test, while the Dumas method is a faster, automated combustion-based alternative for measuring crude protein.
DIAAS over PDCAAS: DIAAS is the updated standard for protein quality, offering more accurate results by measuring ileal (small intestine) digestibility and using uncapped scores, unlike the older PDCAAS method which uses fecal digestibility and caps scores at 1.0.
Colorimetric Assays: Laboratory methods like the Bradford and BCA assays use color changes to quantify protein concentration, but can be sensitive to interfering substances.
Invasive vs. Non-invasive: Methods can range from non-destructive UV spectroscopy to invasive ileal sampling required for precise DIAAS calculation, which impacts cost and practicality.
Choosing a Method: The best method for protein assessment depends on the specific application, considering trade-offs between speed, cost, accuracy, and the required level of detail.

Methods for Protein Quantity Assessment

Assessing protein quantity involves measuring the total amount of protein in a sample, often by determining nitrogen content or using colorimetric assays. The most suitable method depends on factors such as the sample type, required accuracy, and available equipment.

Nitrogen-Based Methods

Kjeldahl Method

Developed in 1883, the Kjeldahl method is a traditional technique for estimating crude protein content by measuring total organic nitrogen. It is often used as a benchmark due to its precision. The process includes digestion with acid and a catalyst to convert nitrogen to ammonium sulfate, followed by distillation into a trapping solution, and finally titration to quantify the nitrogen. A key limitation is that it measures all nitrogen, potentially overestimating true protein, and requires specific or general (like 6.25) conversion factors that can introduce inaccuracies. The method also uses hazardous chemicals and is time-consuming.

Dumas Method

The Dumas method, a modern and automated alternative, determines total nitrogen by combustion. The sample is burned at high temperatures, converting nitrogen to nitrogen gas ($N_2$), which is then measured. This method is faster and avoids hazardous chemicals used in the Kjeldahl method. Like Kjeldahl, it measures non-protein nitrogen and requires a conversion factor to estimate crude protein.

Spectrophotometric Assays

These techniques quantify protein by measuring light absorbance, which changes based on protein concentration.

Bradford Assay

The Bradford assay is a rapid and sensitive method where Coomassie Brilliant Blue G-250 dye changes color upon binding to protein, particularly basic and aromatic amino acid residues. The color change is measured at 595 nm. However, the color response can vary between different proteins and some detergents interfere.

Bicinchoninic Acid (BCA) Assay

The BCA assay is a colorimetric method that is less affected by protein-to-protein variations than the Bradford assay. In an alkaline solution, protein reduces copper(II) ions to copper(I). The BCA reagent then chelates these copper(I) ions, forming a purple complex. The intensity of the purple color, measured at 562 nm, is proportional to the protein concentration.

Methods for Protein Quality Assessment

Protein quality depends on its amino acid composition and digestibility. A high-quality protein provides all essential amino acids in the right amounts and is easily digestible.

Digestibility-Based Scoring Systems

Protein Digestibility Corrected Amino Acid Score (PDCAAS)

Recommended by the FAO/WHO in 1989 and adopted by many regulatory bodies, PDCAAS assesses protein quality by comparing a protein's amino acid profile to a reference pattern based on human needs and correcting for fecal digestibility measured in rats. A significant limitation is that scores are capped at 1.0, meaning proteins with different, potentially superior, profiles can receive the same score. Fecal digestibility can also overestimate absorbed protein due to bacterial activity in the colon.

Digestible Indispensable Amino Acid Score (DIAAS)

Proposed by the FAO in 2013 to replace PDCAAS, DIAAS is an improved scoring system. It uses the digestibility of each individual indispensable amino acid measured at the ileum (the end of the small intestine), providing a more accurate measure of absorption. Unlike PDCAAS, DIAAS scores are not capped, allowing for better ranking of proteins. It also uses updated amino acid requirement patterns for different age groups.

Comparison of Protein Quantity and Quality Assessment Methods


Method	Type of Assessment	Principle	Advantages	Disadvantages
Kjeldahl	Crude Protein (Quantity)	Measures total nitrogen after acid digestion	Widely used, standard for comparison, high precision	Measures non-protein nitrogen, time-consuming, uses hazardous chemicals, not true protein content
Dumas	Crude Protein (Quantity)	Combusts sample to measure nitrogen gas	Faster than Kjeldahl, automated, no hazardous chemicals	Measures non-protein nitrogen, expensive initial setup, not true protein content
Bradford Assay	Protein Concentration (Quantity)	Colorimetric assay based on dye binding	Fast, simple, sensitive	Varies between proteins, susceptible to detergent interference
BCA Assay	Protein Concentration (Quantity)	Colorimetric assay based on copper reduction	Wide linear range, less protein-to-protein variation	Susceptible to interference from reducing agents, reaction is not end-point
PDCAAS	Protein Quality	Corrects amino acid score with fecal digestibility	Standardized by FAO/WHO, widely used for labeling	Scores are capped at 1.0, fecal digestibility overestimates absorption
DIAAS	Protein Quality	Corrects amino acid score with ileal digestibility	More accurate, uncapped scores allow ranking, uses updated reference patterns	More complex, potentially invasive sample collection, not yet widely adopted for regulation

The Evolution of Protein Analysis

The evolution of protein assessment from methods like Kjeldahl and PDCAAS to Dumas and DIAAS reflects a growing need for more accurate nutritional information, especially with the rise of diverse protein sources. This shift towards methodologies that distinguish true protein and accurately reflect human digestibility is important for dietary guidance, food product development, and transparent labeling. While traditional methods still have applications, the trend is towards greater accuracy and specificity.

Conclusion

Understanding how to assess protein quality and quantity is crucial for professionals in food science and nutrition. Quantitative methods like Kjeldahl and Dumas measure total protein, while qualitative systems like PDCAAS and the more accurate DIAAS evaluate nutritional value based on amino acid profile and digestibility. The choice of method depends on the specific analytical goal. The ongoing development of techniques, particularly focusing on ileal digestibility, promises more precise and reliable protein analysis for future nutrition science.

For More Information

For further reading on the evolution of protein quality assessment methods, see this comprehensive review from MDPI: Plant Proteins: Methods of Quality Assessment and the Potential of Plant-Based Proteins for Human Health.

Frequently Asked Questions

Protein quantity refers to the total amount of protein in a sample, often determined by measuring nitrogen content. Protein quality, however, assesses the nutritional value of that protein based on its amino acid composition and how well it is digested and absorbed by the body.

The Kjeldahl method is called a "crude" protein method because it measures total nitrogen, not just the nitrogen found in protein. Non-protein nitrogenous compounds, like melamine, can lead to an overestimation of the true protein content.

PDCAAS stands for Protein Digestibility Corrected Amino Acid Score. It evaluates protein quality by comparing a protein's amino acid profile to a human reference pattern and correcting for fecal digestibility, with scores capped at 1.0.

DIAAS is more accurate because it measures the digestibility of individual amino acids at the ileum (end of the small intestine) rather than the feces. This avoids errors caused by bacterial activity in the colon. DIAAS scores are also not capped, allowing for better ranking of high-quality proteins.

A limiting amino acid is the essential amino acid present in the lowest quantity relative to the body's needs. The protein's overall quality is determined by the concentration of this most limiting amino acid.

These assays quantify protein concentration by measuring changes in color. The Bradford assay uses a dye that shifts color when bound to protein, while the BCA assay uses a reagent that forms a colored complex with copper reduced by the protein.

In jurisdictions like the United States, PDCAAS has traditionally been used for food labeling, but DIAAS is now recommended as the preferred method by the Food and Agriculture Organization (FAO) for its improved accuracy. Some regions also use percentage of total calories from protein for labeling claims.

Yes, food processing can alter protein structure and quality. Heat, for instance, can lead to Maillard reactions that damage essential amino acids like lysine, which can affect the protein's digestibility and its quality score.