**2. Physicochemical properties of fatty acids**

Fatty acids are ubiquitous biological molecules. They are esterified to numerous complex lipid molecules, including triglycerides, phospholipids, and cholesterol esters. As being part of these molecules, fatty acids thus may govern some of their physical properties. The aliphatic chains and their lengths confer hydrophobicity to fatty acids. The hydrophobic nature of the fatty acids renders them insoluble in aqueous environments.

At very high pH, where the longer chained fatty acids are totally ionized, they form micelles, which are thermodynamically stable aggregates of molecules in aqueous solution [21]. This property confers the ionized fatty acids to detergent properties. However, to achieve a stable micelle formation, the fatty acids must be present in a solution at a pH greater than 9, which is generally unphysiologic. In fact, the most probable state of fatty acids at physiological temperature and pH is a membrane-like bilayer structure [22] (**Figure 4**, the middle one). The chain length of the fatty acid is interrelated with melting point; the higher the chain length, the lower the melting point. The double bonds in the (poly)unsaturated fatty acids further decrease their melting points [23]. This is very critical to the survival of mammals that live in extremely cold environments such as the polar areas of the earth. The presence of fatty acids in the bilayer membranes provides an excellent anisotropic solution for other membrane constituents. They confer fluidity to the membrane bilayer [24], wherein membrane-bound receptors, enzymes, and other proteins can diffuse laterally along the surface of the bilayer membrane. Phospholipids can also flip-flop between the bilayer leaflets and/or fatty acyl chains can have a vertical motion (translational motion). The word membrane fluidity can thus be referred to as the degree of stiffness or rigidity of the cellular bilayers. As saturated fatty acids are straight-chained, they can pack/stack easily with themselves and/or with the neighbor-cholesterol in the bilayer membrane. The (poly)unsaturated fatty acyl chains, on the other hand, retain bent(s) along the long axis of the chain at the position of double bonds; thus, they cannot align/stack tightly (**Figure 5**).

Consequently, they increase the degree of membrane fluidity. Therefore, the greater the degree of unsaturation of the fatty acids, the higher the fluidity of the membrane. We have previously reported the DHA, which has six double bonds, contributes to a greater extent in membrane fluidity than less-unsaturated

**Figure 4.**

*The arrangements of fatty acids in aqueous environments at T > Tc. T = temperature. Tc = melting point of the fatty acid.*

#### **Figure 5.**

*The 3D structural features of the most common fatty acids (A). PLA = palmitic acid, STA = stearic acid, OLA = oleic acid, LLA = linoleic acid, LLN = α-linolenic acid, AA = arachidonic acid, EPA = eicosapentaenoic acid, DHA = docosahexaenoic acid. Double bonds of the unsaturated fatty acids are denoted by red color (in A) (B). Because of the presence of double bond(s) along the long axis of (poly)unsaturated fatty acids, they occupy more space when they are esterified in the phospholipid bilayer and loosely align with 3D cholesterol, and increase the degree of disorder (membrane fluidity). However, when straight-chained saturated fatty acids like PLA highly align (stacks) with 3D cholesterol, the degree of packing in the bilayer increases (tightens); hence, the membrane bilayer becomes more rigid, that is, less fluid.*

fattyvacids, such as EPA and/or saturated fatty acids [25, 26]. As a whole, the physicochemical properties of the fatty acids may affect the functions of these molecules [27], ultimately leading to altered functions of the cells and organisms.
