Alkaline phosphatase
[CAS# 9001-78-9]

Identification

• Classification: Food additive >> Enzyme
• Name: Alkaline phosphatase
• Synonyms: 2-[[1-[2-[[1-(2-aminopropanoyl)pyrrolidine-2-carbonyl]amino]acetyl]pyrrolidine-2-carbonyl]amino]-5-(diaminomethylideneamino)pentanoic acid; Alkaline phenyl phosphatase; Alkaline phosphohydrolase; Alkaline phosphomonoesterase; Ostase
• Protein Sequence: APGPR
• Molecular Formula: C₂₁H₃₆N₈O₆
• Molecular Weight: 496.56
• CAS Registry Number: 9001-78-9
• EC Number: 232-631-4
• SMILES: CC(C(=O)N1CCCC1C(=O)NCC(=O)N2CCCC2C(=O)NC(CCCN=C(N)N)C(=O)O)N

Properties

• Density: 1.5±0.1 g/cm³ Calc.^*
• Solubility: water, soluble 0.70 mg/mL (Expl.)
• Index of refraction: 1.682 (Calc.)^*

^* Calculated using Advanced Chemistry Development (ACD/Labs) Software.

Discovory and Applicatios

Alkaline phosphatase is a hydrolase enzyme that catalyzes the removal of phosphate groups from a wide range of phosphorylated substrates under alkaline conditions. It is found in many organisms, from bacteria to humans, and plays essential roles in metabolism, development, and cellular regulation. The enzyme is characterized by its optimal activity at alkaline pH and by its dependence on metal ions such as zinc and magnesium for catalytic function.

The discovery of alkaline phosphatase dates back to the early twentieth century, when researchers studying phosphate metabolism observed enzymatic activities capable of hydrolyzing phosphate esters at elevated pH. Early biochemical experiments identified such activity in animal tissues, particularly in bone, liver, kidney, and intestinal mucosa. As purification techniques improved, alkaline phosphatase was isolated and recognized as a distinct enzyme, separate from acid phosphatases that function optimally at low pH. This distinction helped establish the concept that enzymes with similar catalytic functions could be adapted to different physiological environments.

Structural and mechanistic studies revealed that alkaline phosphatase is a metalloenzyme, with metal ions playing a critical role in stabilizing the active site and facilitating catalysis. The enzyme typically exists as a dimer, with each subunit contributing to catalytic activity. These insights contributed significantly to the development of enzymology, as alkaline phosphatase became a model system for understanding how enzymes achieve specificity and efficiency through metal-assisted catalysis.

In biological systems, alkaline phosphatase serves diverse physiological functions. In humans and other vertebrates, tissue-specific isoenzymes are expressed in bone, liver, intestine, placenta, and other organs. In bone, alkaline phosphatase is closely associated with mineralization, where it contributes to the regulation of phosphate availability during the formation of hydroxyapatite. In the intestine, it is involved in nutrient absorption and detoxification processes. The presence of distinct isoenzymes reflects the enzyme’s adaptation to different biological roles while retaining a common catalytic mechanism.

The measurement of alkaline phosphatase activity has become a routine and important application in clinical diagnostics. Elevated or reduced levels of the enzyme in blood serum are used as indicators of various physiological and pathological conditions, particularly those related to bone metabolism and hepatobiliary function. Because different tissues produce characteristic isoenzymes, analytical methods have been developed to distinguish their origins, enhancing the diagnostic value of alkaline phosphatase assays.

Beyond clinical chemistry, alkaline phosphatase has found widespread application in biotechnology and molecular biology. One of its most influential uses has been as a reporter enzyme in immunoassays and nucleic acid detection systems. Its ability to generate easily detectable signals through the hydrolysis of chromogenic or fluorogenic phosphate substrates made it a cornerstone of early enzyme-linked assays. These applications contributed to advances in diagnostics, research tools, and quality control methods across the life sciences.

In molecular biology laboratories, alkaline phosphatase has also been used as a reagent for dephosphorylating nucleic acids. Treatment of DNA or RNA ends with the enzyme prevents unwanted ligation reactions, facilitating cloning and other genetic manipulations. This practical application highlights how a naturally occurring enzyme was adapted into a precise and reliable laboratory tool.

Alkaline phosphatase from microbial sources has further expanded its utility. Bacterial alkaline phosphatases are often more stable and easier to produce in large quantities, making them suitable for industrial and research applications. Studies of these enzymes have also contributed to comparative biochemistry, revealing how similar catalytic strategies are conserved across species despite differences in sequence and structure.

Overall, alkaline phosphatase represents a classic example of an enzyme whose discovery and characterization have had lasting impact. From its early identification in tissues to its central role in diagnostics, biotechnology, and fundamental enzymology, the enzyme illustrates how detailed study of a single catalytic activity can influence multiple scientific and applied fields. Its continued use and investigation underscore its importance as both a biological catalyst and a versatile research tool.

References

2025. A unique core�shell nanoreactor sSiO2@CeO2/Pt@mSiO2 as artificial nanozyme for ultra-sensitive detection of ascorbic acid and alkaline phosphatase activity. Spectrochimica Acta Part A: Molecular and Biomolecular Spectroscopy.
DOI: 10.1016/j.saa.2025.126459

2025. A novel colorimetric sensor composed from alkaline phosphatase-graphene oxide nanoconjugates and enzyme induced gold nanoparticle aggregation for the detection of deoxynivalenol-3-glucoside. Spectrochimica Acta Part A: Molecular and Biomolecular Spectroscopy.
DOI: 10.1016/j.saa.2025.126433

2025. An electrochemical fluorescence dual-mode strategy for HER2-positive breast cancer cell detection. Talanta.
DOI: 10.1016/j.talanta.2025.127974