The Strait of Malacca (SoM), the world’s busiest sea-route, is increasingly polluted as the rapid development of world trades, affecting phytoplankton primary productivity therein. The variations of surface phytoplankton biomass, size-structure and carbon fixation were investigated across the SoM during the spring period (May 4 to 9, 2011). Chlorophyll a concentration increased from 0.12 µg/L at the northwest entrance of the SoM to a maximal 0.63 µg/L at narrowest section, and decreased to 0.10 µg/L at the southeast entrance. Photosynthetic carbon fixation by phytoplankton coincided well with Chl a biomass, and increased from 10.8 to 22.3 µg C/(L·d), then decreased to 9.21 µg C/(L·d); while the carbon fixation rate showed an inverse pattern to the changes of Chl a, and decreased from 87.1 to 35.5 µgC/(µgChla·d) and increased thereafter to 95.3 µg C/(µg Chl a·d). Picophytoplankton cells (<3 µm) contributed to more than 60% and 50% of the total Chl a and carbon fixation at both the entry waters; while the contributions of pico-cells decreased sharply to the minimum of 18.3% and 27.5% at the narrowest part of the SoM. In particular, our results showed that the silicate concentration positively regulated Chl a biomass and carbon fixation, reflecting that the higher silicate favoured the growth of phytoplankton and thus led to higher primary production in this strait.