Pot experiment was carried out in seventeen levels of fertilization to evaluate the effect of the bio-fertilizer mixture alone or in combination with chemical fertilizer (NPK) on Fennel (Foeniculum vulgare Mill). Bio-fertilizer was a mixture of Azotobacter chroococcum, Bacillus circulans and Vesicular-arbuscular mycorrhiza. Results indicated that use of bio-fertilizer at 3.75 ml/pot four times with 25% of NPK gave the highest significant values of vegetative growth, yield parameters, and also NPK uptake; however, fertilizing with bio-fertilizers without NPK application showed the lowest significant values in the two experimental seasons. The same addition of bio-fertilizer gave the highest significant values of total microbial count of soil and increased cumulative CO2; whereas addition of 100% NPK gave the lowest significant values in both seasons. The full dose of NPK significantly increased volatile oil percentage while the addition of bio-fertilizer only significantly decreased it in both seasons. GC analysis of essential oil showed that the highest percentage of anethol (12.77 %) was obtained by applying 3.75 ml/pot bio-fertilizer added one time + 25% NPK and decreased estragole (72.78%).