Dairy Chemistry & Milk Constituents

Chemical origin of heated milk flavours:

• Butyric acid is a short-chain free fatty acid liberated by the hydrolytic action of lipoprotein lipases; it produces a sharp, pungent rancid off-flavor (lipolyzed/hydrolytic rancidity), NOT a cooked flavor.
• Volatile sulphur compounds (hydrogen sulphide, methanethiol, dimethyl sulphide) generated by thermal denaturation of $\beta$-lactoglobulin sulfhydryl groups are the primary cause of cooked / cabbagey flavor.
• Lactones ($\delta$-decalactone, $\delta$-dodecalactone) generated from hydroxy fatty acids contribute sweet, peachy cooked notes.
• Methyl ketones generated by thermal decarboxylation of $\beta$-keto fatty acids contribute rich heated/stewed milk flavors.

Thermodynamic heat stability and denaturation kinetics of whey proteins:
When measured by irreversible loss of solubility upon heat treatment:

• Immunoglobulins (B): Most heat-sensitive bovine whey protein; rapidly denatures and aggregates irreversibly between $60\text{--}65^\circ\text{C}$ (lowest heat stability).
• Bovine Serum Albumin, BSA (C): Denatures at $64\text{--}68^\circ\text{C}$.
• $\beta$-Lactoglobulin (D): Denatures at $70\text{--}78^\circ\text{C}$, exposing its free cysteine thiol group and forming irreversible intermolecular aggregates.
• $\alpha$-Lactalbumin (A): Possesses an apparent unfolding temperature around $62\text{--}65^\circ\text{C}$, but because its four disulfide bonds and bound $Ca^{2+}$ ion allow remarkable reversible renaturation upon cooling, it resists permanent loss of solubility better than all other major whey proteins (highest heat stability).

Pigment distribution in milk phases:

• Carotene is a fat-soluble pigment imparting golden yellow color to milk fat and cow butter, but partitions into fat globules, not liquid whey.
• Anthocyanin is a water-soluble plant pigment occurring in berries and flowers.
• Chlorophyll is the green photosynthetic plant pigment.
• Riboflavin (Vitamin $B_2$) is a water-soluble vitamin naturally dissolved in the aqueous serum phase of milk (~$1.5\text{ mg/L}$). During cheese or paneer making, when fat and casein are removed as curd, the unobstructed fluorescence of dissolved riboflavin imparts the characteristic distinctive greenish-yellow tint to liquid whey.

Polymorphism of bovine $\beta$-casein at codon 67:
(A), (C), (D) misassign the amino acid residues.

• The distinction between $A^1$ and $A^2$ beta-casein arises from a single nucleotide polymorphism (SNP, CCT $\rightarrow$ CAT) at codon 67 of the $\beta$-casein gene:
• $A^2$ variant (the ancestral wild-type allele found predominantly in Bos indicus zebu cattle): Possesses Proline at position 67 ($A^2\text{ - Proline}$). Because of the rigid cyclic pyrrolidine ring of proline, the adjacent peptide bond between residues 66 and 67 resists enzymatic cleavage.
• $A^1$ variant (a mutation originating in European Bos taurus cattle): Possesses Histidine at position 67 ($A^1\text{ - Histidine}$). Gastrointestinal digestive proteases (pancreatic elastase) readily cleave the peptide bond adjacent to histidine, liberating the 7-amino acid bioactive opioid peptide beta-casomorphin-7 (BCM-7).