Human milk is recommended as the optimal source of nutrition for babies up to six months of age, as it provides individualized, specific and abundant nutritional compounds, as well as immunological protection. Currently, there is wide interest in studying human milk because of this key role, as well as identification of biologically active compounds. In addition, human milk banks play an important role in supplying donor human milk to infants, especially premature infants, in hospitals, which involves processing and storage of such milk. This review summarises recent studies on macronutrients in human milk, including their structure, levels, influencing factors, and contributions to the health benefits of infants. Physico-chemical properties of human milk are also reviewed, including pH, colour, stability of casein micelles, size and zeta-potential of fat globules, heat stability, freeze-thaw effect, dehydration and phase separation, and the impact of preservation processes on these. The levels of human milk macronutrients and factors that influence these, such as lactation and gestational age, are ongoing research topics. Studies on physicochemical properties of some human milk components, such as caseins, are comparatively few, as research has mainly focused on bioactive peptides, milk fat globule membrane and human milk oligosaccharides. Processing and preservation are applied to human milk and the effects of these methods on the retention of human milk nutrients for use in human milk banks and hospitals must be considered. Some physico-chemical properties, such as milk pH, colour, and physical stability, may be possible indicators of milk quality. •Studies on the levels of macronutrients in human milk are summarized.•Physiochemical properties of human milk are reviewed.•Technologies for preservation of human milk, such as heating, high-pressure treatment and drying, are discussed.•Reports on the stability of human milk nutrients following such preservation steps are reviewed.