We have synthesized first novel dihexadecyl phosphate (DHGP-n) gemini surfactants containing alkyl chain ((C n H 2n ); n = 3, 4, 5, and 6) spacers. Surfactants were used to form monolayers on water surface. Surface tension measurement (STm), Brewster angle microscopy (BAM), and density functional theory (DFT) calculation were used to investigate these monolayers. Surface pressure (π) and molecular area (A); π-A isotherm curves of all DHGP-n gemini surfactants showed a gradual increase in

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... pressure without any break points in the curves, confirmed the formation of liquid-expand (LE) type monolayers. BAM observations also supported the STm results. Limited molecular area (A 0 ) of these monolayers depended on the spacer of gemini surfactants without spacer n = 4. Longer alkyl chain spacer in the series had higher value of A 0 in π-A isotherm plots. In the series, A 0 of spacer n = 4 was smallest among A 0 values of other spacers. Density functional theory (DFT) method calculation also confirmed π-A isotherm curves pattern of these surfactants. Calculation showed that both odd and even numbers and length of the alkyl chain spacer influenced the structure of DHGP-n monolayer formation on the water surface.