The Impact of Carotenoid Energy Levels on the Exciton Dynamics and Singlet-Triplet Annihilation in the Bacterial Light-Harvesting 2 Complex

The light-harvesting 2 (LH2) complex of purple phototrophic bacteria plays a critical role in absorbing solar energy and distributing excitation energy. Exciton dynamics within LH2 complexes are controlled by the structural arrangement and energy levels of the bacteriochlorophyll (BChl) and carotenoid (Car) pigments. However, there is still debate over the competing light-harvesting versus energy-dissipation pathways. In this work, we compared five variants of the LH2 complex from genetically modified strains of Rhodobacter sphaeroides , all containing the same BChls but different Cars with increasing conjugation: zeta-carotene ( N =7; LH2 _Zeta ), neurosporene ( N =9; LH2 _Neu ), spheroidene ( N =10; LH2 _Spher ), lycopene ( N =11; LH2 _Lyco ), and spirilloxanthin ( N =13; LH2 _Spir ). Absorption measurements confirmed that Car excited state energy decreased with increasing conjugation. Similarly, fluorescence spectra showed that the B850 BChl emission peak had an increasing red shift from LH2 _Zeta →(LH2 _Neu /LH2 _Spher )→LH2 _Lyco →LH2 _Spir . In contrast, time-resolved fluorescence and ultrafast transient absorption (fs-TA) revealed similar excited state lifetimes (∼1 ns) for all complexes except LH2 _Spir (∼0.7 ns). From fs-TA analysis, an additional ∼7 ps non-radiative dissipation step from B850 BChl was observed for LH2 _Zeta . Further, singlet-singlet and singlet-triplet annihilation studies showed a ∼50% average fluorescence lifetime reduction in LH2 _Zeta at high laser power and high repetition rate, compared to ∼10-15% reductions in LH2 _Neu /LH2 _Spher /LH2 _Lyco and minimal lifetime change in LH2 _Spir . In LH2 _Zeta , the fastest decay component (<50 ps) became prominent at high repetition rates, consistent with strong singlet-triplet annihilation. Nanosecond TA measurements revealed long-lived (>40 μs) BChl triplet states in LH2 _Zeta and signs of damage caused by singlet oxygen, whereas other LH2s showed faster triplet quenching (∼18 ns) by Cars. These findings highlight a key design principle of LH2 complexes: the Car triplet energy must be significantly lower than the BChl triplet energy to efficiently quench BChl triplets that otherwise act as potent “trap states” causing exciton annihilation in laser-based experiments or photo-damage in native membranes.