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[en] Highlights: • Polyynes were synthesized by femtosecond laser ablation of SWCNTs in methanol. • SERS spectra of isolated polyynes in Ag colloid were investigated. • Clear peaks in the region of β band of C8H2 were observed in the damp-dried colloid. Polyyne samples C2nH2 (n = 4–6) were synthesized by irradiating single-wall carbon nanotubes in methanol with intense femtosecond laser pulses. For obtaining isolated polyynes (C8H2, C10H2, and C12H2), the original solution was separated by high performance liquid chromatography. The surface-enhanced Raman scattering spectra of isolated polyynes in Ag colloid have been investigated with naturally drying time, and clear peaks in the region of β band for the isolated C8H2 were observed at 1910 and 1958 cm−1 in the damp-dried Ag colloid samples for the first time.