Biochemical studies on blackspot bruising in potato (Solanum tuberosum L.)
Potatoes respond to mechanical stress by invoking a precisely controlled biochemical reaction leading to production of the pigment melanin. Predisposition to produce melanin is variety specific, this specificity being conveyed by a range of genetic, environmental and biochemical factors. A well characterised response of plant tissues to perturbation by biotic or abiotic stress factors is the rapid generation of oxygen based free radicals - active oxygen species (AOS). Due to their nature as high-energy, largely unstable, radicals, AOS have been implicated in a wide range of deleterious effects upon plant tissues as well as being involved in a range of signalling cascades within cells. This project attempted to demonstrate a link between AOS and other associated oxidative factors, and blackspot bruise potential. It was demonstrated that in response to mechanical impact, potato tubers respond by producing a well-defined level of superoxide free radicals. The amount of superoxide generation was found to be nearly perfectly correlated with susceptibility to blackspot bruise when investigated over eight potato varieties. By using inhibitors and scavengers of free radicals it was shovra that superoxide and not hydrogen peroxide was directly responsible for the effects upon pigment synthesis. Furthermore studies upon polyphenol oxidase (PPO), the key enzyme in the synthesis of melanin, demonstrated that a significant enhancement of activity was noted in the presence of superoxide radicals, lending support to animal studies which suggest that PPO utilizes superoxide radicals more readily and more efficiently than its usual co-substrate molecular oxygen. Associated with the production of superoxide radicals was the demonstration of modification to tuber proteins, specifically the detection of secondary carbonyl groups, a modification known to be the direct result of exposure to AOS. This study quantified the carbonyl levels and once again these were almost perfectly correlated with susceptibility to blackspot bruise formation. Once the correlation of oxidative factors and blackspot bruise susceptibility was established then studies were undertaken to demonstrate the effect of a range of metalions upon both blackspot bruise susceptibility and oxidative protein modifications. These studies suggested that potassium and zinc both had negative effects on blackspot susceptibility however only potassium had a negative effect on secondary carbonyl accumulation. Studies on tubers impacted twice indicated that the normal 'polarity' of the tuber in terms of susceptibility to blackspot bruise could be influenced and this was confirmed by studies on superoxide generation which showed that the second point of impact always had a reduced blackspot susceptibility and reduced superoxide generation. Finally a model is presented to incorporate this novel information regarding the role of AOS in blackspot bruise susceptibility together with existing knowledge of the biochemistry within this system, to propose a system in which AOS play an integral and wide-ranging role.