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What happens to donkey milk whey protein when it is broken down into smaller pieces using enzymes? A team from Liaoning University in China compared four common laboratory enzymes and measured the resulting mixtures (hydrolysates) in the lab. The broken-down samples were more soluble and had smaller particles than the original protein. However, the paper contains several contradictions between the text and the tables.
Food technology laboratory study (in vitro) – physical and chemical properties were measured in test tubes; it makes no statement about effects in humans. Donkey milk is a food.
Study at a glance
| Feature | Information |
|---|---|
| Original title | Characterisation and comparison of enzymatically prepared donkey milk whey protein hydrolysates |
| Authors | Wei M., Ning C., Ren Y. et al. |
| Journal | Food Chemistry: X, 22: 101360 |
| Year | 2024 |
| Country | China |
| Study type | Food technology laboratory study (in vitro) |
| Sample | Purchased donkey milk whey protein; five samples (original protein and four hydrolysates), each measured in triplicate |
| DOI | 10.1016/j.fochx.2024.101360 |
What question did the researchers ask?
Whey proteins are the proteins that remain in the whey when casein is separated. The team wanted to know how four enzymes – subtilisin, bacillolysin, papain and a peptidase mix – affect the structure and technical properties of donkey milk whey protein: solubility, particle size, emulsification and foaming behavior, as well as values in chemical radical tests.
How it was investigated
- Preparation: A 6% solution of the whey protein was heated for 10 minutes at 85 °C.
- Breakdown: One enzyme each at a ratio of 4:100 to the protein, for 4 hours (subtilisin at pH 9.0 and 55 °C, the others at pH 7.0 and 50 °C), with ultrasound for the first 20 minutes.
- Processing: Enzymes deactivated by boiling, centrifuged, supernatant freeze-dried.
- Measurement: Degree of hydrolysis (proportion of broken bonds), molecular size in gel, protein folding, amino acids, solubility, particle size, emulsion, foam and three radical tests.
The results
The peptidase mix broke down the most (degree of hydrolysis 39.46%), followed by bacillolysin (30.36%) and papain (12.73%); the text does not give a numerical value for subtilisin. All hydrolysates were more soluble and had smaller particles than the original protein.
| Measurement | Whey protein | Hydrolysates |
|---|---|---|
| Solubility | 47.74 % | Subtilisin 71.11 %; others "close to 80 %" |
| Mean particle size | 309.57 ± 11.32 nm | 214.57 ± 4.26 nm (bacillolysin) to 261.97 ± 19.67 nm (subtilisin) |
| Emulsion stability | 61.62 ± 0.20 min | Bacillolysin 75.94 ± 0.34 min; peptidase mix 93.05 ± 1.45 min |
| Total amino acids | 178.68 mg/g | 289.62 to 320.95 mg/g |
In the radical tests, the paper only gives figures for the peptidase mix hydrolysate at 2.5 mg/mL: 59.00 ± 0.58 % (DPPH), 52.97 ± 1.55 % (hydroxyl radical) and 44.50 ± 0.48 % (ABTS). The values for the other samples are only in one figure.
The choice of enzyme changed how strongly the donkey milk whey protein was broken down and how the mixtures behaved in the lab. These are food technology laboratory measurements.
Critical assessment
- Contradictions: The text speaks of "identical amino acid composition", while the table shows clear differences (glycine 14.66 mg/g compared to around 5 mg/g). The folding measurement is interpreted as a loss of α-helix, but according to the table, it increased in three out of four hydrolysates.
- Molecular size: The text mentions peptides under 5 kDa, but bands around 25 and 23 kDa are still visible in the gel for two enzymes.
- Missing figures: No degree of hydrolysis for subtilisin, no figures for emulsification and foaming capacity in the text, radical tests quantified for only one sample.
- Unknown milk: The whey protein was purchased; the origin and production of the donkey milk are not described.
- Conclusion on the data: The authors view the hydrolysates as an antioxidant food additive. Intake or application were not investigated; they themselves call their radical tests "not comprehensive enough".
Funding, conflicts of interest and level of evidence
The work was funded by the Liaoning Provincial Natural Science Foundation (2022-BS-112). The authors declare no conflicts of interest. The whey protein comes from a Chinese biotech company.
Level of evidence: Food technology basic research in the lab (in vitro) – describes properties of protein mixtures, makes no statement about effects.
What does this mean for donkey milk as a food?
The study describes how donkey milk whey protein changes technically when it is specifically broken down with enzymes. This laboratory process has little to do with donkey milk as it is drunk as a food. We do not adopt the authors' ideas for application.
Our research question: How strongly does even heating change donkey milk whey proteins – and how do they react to freezing or drying?
Frequently asked questions
What is a hydrolysate?
A mixture of protein fragments that is created when protein is broken down with enzymes or acid. The degree of hydrolysis indicates how far the breakdown has progressed.
Which enzyme came out on top?
It depends on the measurement. The peptidase mix achieved the highest degree of hydrolysis and the longest emulsion stability, bacillolysin the smallest particles. A ranking for everyday life cannot be derived from this.
Was anything tested on humans or animals?
No. All measurements took place in test tubes.
You can find all measurements, interactive graphics and the complete critical assessment on our research page: Study 81: Donkey milk whey protein, broken down with four enzymes. Further studies in the research archive.
Wei M., Ning C., Ren Y., Hu F., Wang M., Li W. (2024). Characterisation and comparison of enzymatically prepared donkey milk whey protein hydrolysates. Food Chemistry: X, 22, 101360. DOI: 10.1016/j.fochx.2024.101360
Summarized and checked by the DON DONKEY'S editorial team based on the original publication · As of: October 2026. This summary does not replace the original work.
End of study summary. The study was conducted independently of DON DONKEY'S® and does not refer to our products. Our donkey milk is a food.
