In order to improve the extraction ability of carbon fibers (CFs) for microextraction ofpolycyclic aromatic hydrocarbons (PAHs), biochar nanospheres derived from glucose were in-situ grown onto the surface of CFs via hydrothermal synthesis. The surface morphology and elemental composition of biochar nanospheres-CFs were investigated by scanning electron microscopy and X-ray photoelectron spectroscopy. Thereafter, the biochar nanosphere-CFs were pulled into the polyetheretherketone tube for solid-phase microextraction, and the tube was combined with high-performance liquid chromatography-diode array detector to online detect PAHs. With the help of π-stacking, hydrophobic, and hydrophilic effect of biochar nanospheres, the extraction efficiency of CFs was greatly enhanced (enrichment factor increased by 293% compared with the original). The conditions affecting the analytical performance (sampling volume, sampling rate, methanol content, and desorption time) were investigated. Under the optimal conditions, an online analytical method for microextraction and determination ofseveral PAHs was developed, and satisfactory results were achieved. The limits of detection were 0.003-0.010 ng mL-1 owing to high enrichment effect (2973-3600), linearity ranged from 0.010-15.0 ng mL-1, andrelative standard deviations were 0.4%-1.6% (intra-day) and 2.4%-4.4% (inter-day), respectively. The method was applied toanalyze environmental water samples (rain water, snow water, and river water), and spiked recoveries within 80.0%-119% were obtained.