The aim of this paper is to estimate the impact of changing channel spacing in a wavelength division multiplexed (WDM) transmission system on the performance of the in-line erbium doped fiber amplifier (EDFA). The performance of the EDFA was investigated in a 16 channel 9.953 Gbit/s WDM transmission system with non-return-to-zero encoding technique, intensity on-off keying modulation format (NRZ-OOK) and three different values of channel spacing (37.5 GHz, 50 GHz and 100 GHz). The single-pump EDFA solution with co-propagating pumping radiation and 12 m long erbium doped fiber was used. To estimate the impact of changes of channel spacing on the performance of the in-line EDFA, the minimal pump power that is required to ensure a certain quality of the signal for each channel spacing was found. To assess the performance of the EDFA, the dependences of the maximal BER values among all the channels on the power of the detected signals for each channel spacing are compared to the same results obtained in a 16 channel WDM system with the corresponding channel spacing but without amplification. Power penalty values are evaluated at each channel spacing in respect to the solution without amplification.