Bacteria can use n-hexadecane as a carbon source, but it remains incompletely understood whether n-hexadecane is transformed into metabolic intermediates prior to cellular uptake or not. We newly isolated a strain identified as Pseudomonas synxantha LSH-7' and conducted chemotaxis experiment of this bacterial strain towards n-hexadecane, hexadecanol and hexadecanoic acid with qualitative assays respectively. Furthermore, we described the identification of extracellular alkane hydroxylase and alcohol dehydrogenase activity; acidification of the culture medium; identification of hexadecanoic acid in the culture medium by the GC-MS analysis; and variation concentration of intracellular n-hexadecane and hexadecanoic acid. A detailed analysis of the experimental data revealed the chemotaxis of this bacterial strain towards n-hexadecane instead of its metabolic intermediates. Our results further suggested that only a fraction of total n-hexadecane followed this path, and alkane hydrolase and hexadecanol dehydrogenase were constitutively expressed when grown in the medium of n-hexadecane. Most strikingly, we quantitatively investigated the concentration of n-hexadecane adsorbed by bacterial chemotaxis. Our findings provided an original insight n-hexadecane might be converted to hexadecanoic acid extracellularly before it was taken up across the cell membrane.