We present a comprehensive experimental study of protonated tyramine ions in a cold 3D quadrupole ion trap coupled to a time-of-flight mass spectrometer. Multiple UV photodissociation techniques have been developed, including single and double resonance spectroscopy along with time-resolved excited state lifetime measurements through a picosecond pump–probe scheme. An original UV–UV hole burning method is presented which can be used without modification of the quadrupole ion trap. The electronic spectrum of the cold protonated tyramine exhibits well-defined vibronic transitions, allowing the firm assignment of its two low-lying energy conformations by comparison with CC2 ab initio excited state calculations.