Two of the most familiar additives in the dairy aisle, sodium dihydrogen phosphate and disodium hydrogen phosphate, have long been treated by food technologists as quiet workhorses: emulsifying salts, pH regulators and stabilizers that keep processed cheese smooth and shelf-stable. A new study published in Food Chemistry: X suggests they are anything but passive. In a detailed model of the Maillard reaction, the non-enzymatic browning process that unfolds when proteins meet reducing sugars under heat, the two phosphate salts drove casein glycation in strikingly different directions, one accelerating browning and flavor-generating chemistry, the other restraining it. The findings could reshape how manufacturers control color, texture and flavor stability in heat-treated dairy products.

The research team, led by Shaohong Xu and colleagues, built a simplified but rigorous experimental system: casein, the predominant protein in milk, was mixed with glucose at a fixed concentration and incubated at 90 degrees Celsius for up to 24 hours. Into this mixture they added either sodium dihydrogen phosphate, abbreviated MSP, or disodium hydrogen phosphate, DSP, at low and high doses of 10 and 100 millimolar. Crucially, the pH of every sample was adjusted to 7.0 before the reaction began, a design choice that allowed the researchers to isolate the intrinsic chemical effects of the phosphate species and ionic strength from the well-known influence of acidity itself. Samples were then pulled at eight time points, from half an hour to a full day, and interrogated with an arsenal of analytical techniques.

The most visually dramatic result came from simple colorimetry. After 24 hours of heating, the browning index of the untreated control casein reached about 28 units. Adding low-dose MSP slashed that figure to roughly 4.5, producing a nearly white sample, while high-dose MSP still kept browning well below the control at about 17 units. DSP did the opposite: low-dose DSP pushed the index to about 41, and high-dose DSP sent it soaring to nearly 222, an eightfold increase over the control, accompanied by a deep reddish-brown hue. The explanation lies in acid-base chemistry. MSP is a weakly acidic phosphate that pulls the local reaction environment toward lower pH values unfavorable to the Maillard cascade, whereas DSP, a weakly alkaline salt, nudges the local pH upward and thereby facilitates the initial attack of glucose on the protein’s amino groups.

Spectroscopic measurements traced the reaction’s progress through its intermediate and advanced stages. Ultraviolet absorbance at 294 nanometers, a proxy for early Maillard intermediates such as furfurals and reductones, rose steadily in all samples over 24 hours, but the 100 millimolar phosphate groups, both MSP and DSP, accumulated significantly more intermediates than the control throughout the run. Absorbance at 420 nanometers, which tracks the colored melanoidin pigments of the reaction’s late stage, told a complementary story: every phosphate-supplemented sample browned faster than the control, and the effect was clearly concentration-dependent, with the high-dose groups reaching the highest pigment levels. Even the low-dose groups, though weaker, remained measurably above the control, hinting that ionic strength itself can redirect reaction pathways and blunt pigment formation efficiency.

Beneath the color changes, the phosphates were quietly remodeling the protein itself. Dynamic light scattering revealed that the untreated control casein underwent dramatic particle growth during glycation, a signature of aggregation and conformational unfolding. Phosphate-treated samples, by contrast, showed far gentler increases in particle size, and the effect scaled with dose: 100 millimolar phosphate consistently yielded smaller particles than 10 millimolar. This mirrors earlier observations in meat and dairy systems, where phosphates reduce protein diameter and boost solubility through charge shielding and dispersion mechanisms. In practical terms, phosphate addition kept the glycated protein dispersed rather than clumped, a property that also improved water solubility and emulsifying capacity, traits highly prized in beverage and encapsulation applications.

The salts also unmasked the protein’s reactive chemistry. Using the OPA assay to quantify free amino groups, the researchers found that at the very start of the reaction, before any glycation had proceeded, all phosphate-treated samples displayed significantly higher free amino content than the control, with the high-dose DSP group recording the highest value. This indicates that phosphate pretreatment exposes lysine and arginine residues that are normally buried inside the folded protein, handing the glucose molecule more accessible docking sites. Over the first two hours, the phosphate groups showed dynamic shifts, with some samples consuming free amino groups early while the high-dose MSP group sustained elevated levels, demonstrating persistent reactive capacity. Phosphate, in other words, acts not merely as a dispersant but as a molecular chaperone of sorts, rearranging the protein to make it more reactive.

Circular dichroism spectroscopy confirmed that this rearrangement was structural and remarkably fast. Far-UV spectra collected at one, two and 16 hours showed that glycated samples shifted from a highly ordered, beta-sheet-rich conformation toward a more flexible, disordered state. Alpha-helix content rose modestly from about 6 percent in the control to 8 or 9 percent in treated samples, while parallel beta-sheet structure plummeted by 10 to 15 percentage points, and beta-turn and random coil fractions climbed correspondingly. Strikingly, the secondary structure proportions changed by less than 2 percent across the three time points within each sample, meaning the major conformational overhaul was essentially complete within the first hour of heating and then held steady. The loosened, open architecture of the high-dose phosphate samples aligns neatly with their higher free amino exposure and faster glycation kinetics.

To map exactly where glucose and its reactive descendants attached, the team turned to proteomics, digesting the glycated casein with trypsin and analyzing the resulting peptides by nano-scale liquid chromatography coupled to an Orbitrap mass spectrometer. They catalogued glycation sites, carboxymethylation, carboxyethylation, pyrraline, and the hydroimidazolone derivatives of glyoxal and methylglyoxal on lysine and arginine residues. After one hour, the MSP-treated samples showed markedly fewer detectable modification sites than the control, particularly for glucosylation and oxidation, while the DSP samples carried as many or more sites, especially for carboxymethylation and carboxyethylation. By four hours the picture shifted: MSP’s site counts rose steeply, suggesting its early inhibitory grip weakens with time, whereas some DSP markers declined, implying dynamic conversion or masking of modified residues. The authors caution that these counts reflect detectable site distribution rather than absolute glycation extent, since the analysis was not based on peptide-level quantification.

Perhaps the most commercially consequential findings concern flavor. An electronic nose fitted with ten sensors for compound classes ranging from aromatics to sulfides showed that after one hour, the control and high-dose DSP samples emitted the strongest volatile signals, while the MSP groups, especially the low-dose one, yielded the weakest. Gas chromatography-ion mobility spectrometry sharpened the picture: the untreated sample reached normalized intensity values of 100 for aldehydes and alcohols such as hexanal and 1-hexanol, evidence of amino acid degradation and Strecker degradation pathways, whereas high-dose MSP dramatically suppressed pentanal, 2-heptanone and ethyl propanoate. DSP-treated samples, meanwhile, maintained aldehyde, ketone and ester signals comparable to or above the control. Because the model system contained no lipids, these volatiles arose purely from protein-side chemistry, making the contrast especially clean.

Taken together, the study recasts phosphate salts as active, selective regulators of the Maillard reaction rather than inert processing aids. At high concentrations, both salts accelerate glycation by curbing aggregation, exposing amino groups and loosening protein structure, yet they diverge sharply downstream: MSP restrains early glycation and oxidative modification and suppresses aldehyde and ketone formation, supporting flavor stability, while DSP promotes browning and advanced glycation end product accumulation with little effect on the volatile profile. For manufacturers of processed cheese, milk powders and heat-treated dairy formulations, the message is that the choice between these two chemically similar salts is a choice between two different chemical futures for the product, one browner and more aromatically active, the other paler and more flavor-stable. The work offers a mechanistic roadmap for exploiting that divergence deliberately, turning a longstanding empirical art of dairy formulation into a matter of precise molecular control.

Subject of Research: Effects of sodium phosphate salts on the Maillard glycation of casein in dairy systems

Article Title: Effects of sodium dihydrogen phosphate (MSP) and disodium hydrogen phosphate (DSP) on the glycation of casein (CN)

Article References: Xu, S., Hu, X., Sun, Y., Huang, X., Zhou, Z., Han, L., Zhang, Y., Zhang, J., Gao, Y., Wang, L., & Qin, D. (2026). Effects of sodium dihydrogen phosphate (MSP) and disodium hydrogen phosphate (DSP) on the glycation of casein (CN). Food Chemistry: X, 39, Article 104621. https://doi.org/10.1016/j.fochx.2026.104621

Image Credits: AI Generated

DOI: 10.1016/j.fochx.2026.104621

Keywords: Maillard reaction, casein, glycation, phosphate salts, dairy processing, browning, food chemistry, protein structure, volatile compounds, flavor stability, processed cheese, mass spectrometry