The susceptibility of the 'Pinkerton' avocado cultivar to mesocarp discolouration, after storage, has seriously threatened its export from South Africa. This disorder has proven to be complex, requiring a better understanding of the fruit's physiology. The purposes of this study were to identify the role of pre- and postharvest factors, or their interactions, in the development of the problem. This was done by obtaining fruit from several production areas of varying mesocarp discolouration histories (referred to as "high", "medium" or "Iow risk" areas) during the 2000 and 2001 seasons. Fruit were stored at 8, 5.5 and 2°C for 30 days, as well as ambient (20°C). Evaluations of fruit quality were made before and after storage, as well as after softening. Once removed from storage the weight loss (during storage) was determined, and fruit firmness and carbon dioxide (C02) production rates monitored daily. It was found that temperatures below the recommended shipping temperature of 5.5°C, i.e. 2°C, produced the best internal fruit quality. This was supported by the membrane integrity studies that showed less membrane stability at the warmer storage temperature of 8°C. Furthermore, remained hard during storage and subsequently had an extended shelf life. Fruit origin was also found to play a major role in browning potential, with discolouration being consistently more severe in fruit from "high risk" areas and increasing in severity as the season progressed. The rate of CO2 production was found to follow a similar trend, with rates increasing as the season progressed, and also being slightly higher in fruit from "high risk" areas. The higher CO2 production rates were thought to be related to a decrease in membrane integrity as the season progressed. While storage temperature was not found to have a significant effect on the rate of CO2 production after storage, it did affect the time taken to reach the maximum rate, with fruit stored at 2°C taking longer. Biochemical analyses to determine the concentration of total phenolics and the activity of the enzyme polyphenol oxidase (PPO) also showed that the potential for browning was initiated by preharvest conditions. While no significant differences were found between growers with regards to total phenol concentrations, the PPO activity was found to be higher in fruit from poor quality areas, and subsequently browning potential was expected to be higher in these fruit. It was, however, found that the potential for browning could be reduced by storing fruit at 2°C, as this decreased the total phenolics concentration. This evidence further emphasized the idea that storage at 2°C could be highly advantageous. Fruit mineral analysis showed that certain key elements played a significant role in the severity of mesocarp discolouration, with excessive fruit nitrogen and decreasing copper and manganese concentrations appearing to play major roles. The high fruit nitrogen concentrations were suspected to reflect fruit grown on very vigorous trees, resulting in shoots competing with fruit for available reserves. It is suggested that 'Pinkerton' of a quality acceptable to the market, can be produced by manipulating source:sink relationships, particularly through decreasing the availability of nitrogen, followed by low temperature (24° C) shipping. Future work should concentrate on manipulation of source:sink relationships, to take account of both climatic conditions and leaf to fruit ratios. The evaluation of chlorophyll fluorescence as a tool for predicting mesocarp discolouration potential in 'Pinkerton' proved to be unsuccessful in this study and future studies may require modifications to the current technique. It is suspected that differences in chlorophyll content, for example, between fruit from different origins, will have to be taken into account when interpreting results. The success of using 2°C storage to improve the internal quality on 'Pinkerton' fruit prompted further studies, during 2004, to ensure that the development of external chilling injury would not decrease the marketability of the cultivar. Low temperature conditioning treatments, prior to storage, proved to be highly successful in reducing the development of external chilling injury, thus further improving fruit quality as a whole. Preconditioning treatments consisted of fruit that were kept at either 10°C, 15°C or 20°C for 1 or 2 days before being placed into storage for 30 days at 2°C or 5.5°C. All preconditioning treatments were compared to fruit that were placed directly into storage. The effect of fruit packaging on moisture loss (as determined by weight loss) and chilling injury was also investigated using unwaxed fruit, commercially waxed and unwaxed fruit individually sealed in micro-perforated polypropylene bags with an anti-mist coating on the inside (polybags). Holding 'Pinkerton' fruit, regardless of packaging treatment, at 10°C for 2 days prior to storage at 2°C or 5.5°C significantly decreased the severity of external chilling injury. The use of polybags during preconditioning and storage showed potential in further reducing the development of external chilling injury, although the higher incidence of fungal infections in these fruit needs to be addressed. The determination of proline concentrations in fruit exocarp tissue after storage was helpful in determining the level of stress experienced by fruit that were subjected to different packaging and preconditioning treatments. In this study waxed fruit subjected to 1 d preconditioning at 10°C, 15°C or 20°C or placed directly into storage at 2°C showed very high proline concentrations and also displayed more severe external chilling injury, despite unwaxed fruit losing more weight during these treatments. The role of moisture loss thus needs further investigation. The thickness and method of wax application was thought to play an important role in the higher external chilling injury ratings in this study as waxed fruit often developed chilling injury symptoms around the lenticels and it was suspected that either the lenticels were damaged by the brushes used to apply the wax or that the lenticels became clogged thus resulting in reduced gaseous exchange. Nevertheless, the success of low temperature conditioning in reducing external chilling injury, while maintaining sound internal quality, may enable storage temperatures to be dropped even further, thus enabling South Africa to export avocados to countries that require a cold disinfestation period prior to entry to eliminate quarantine pests (e.g. fruit fly). / Thesis (Ph.D.Agric.)-University of KwaZulu-Natal, Pietermaritzburg, 2005.

New emerging markets such as Japan and the United States require cold sterilisation of South African citrus fruit as a phytosanitary standard against fruit fly. However, citrus fruit are chilling susceptible, with lemons being the second-most chilling susceptible after grapefruit. Chilling injury is a physiological rind disorder; the occurrence of which is despite its prevalence in horticultural commodities, not well understood. Therefore, the aim of this study was to investigate physiological compounds regulating chilling susceptibility or resistance in citrus fruit, with special emphasis on lemons. Furthermore, the potential of hot water dips or “molybdenum soaks” to maintain a certain level of physiological compounds which determine manifestation of chilling injury symptoms in citrus fruit was investigated. Moreover, it was attempted to create an understanding of the order in which physiological compounds mitigate chilling injury. Lemon fruit from different farms known to be chilling susceptible or resistant were obtained during the 2007 and 2008 harvest season. Thereafter, fruit were treated by soaking for 30 min in 1μM NaMo04.2H20 solution followed by a 2 min HWD 47 or 53°C. Treated fruit were waxed, weighed and stored at -0.5°C for up to 28 days and sampled for chilling injury evaluation 7, 14, 21, or 28 days into cold storage. A second evaluation was carried out five days after withdrawal from cold storage to allow development of chilling injury symptoms as a shelf-life simulation. After the second evaluation fruit were peeled, peel freeze-dried, milled using mortar and pestle and stored at -21°C for further physiological analysis. Freeze-dried peel was analysed for soluble sugars (glucose, fructose, sucrose), vitamin C (ascorbic acid), vitamin E (α-tocopherol), β-carotene, polyamines (putrescine, spermine, spermidine), specific flavanones (naringin and hesperidin) using HPLC-UV-Vis detector and proline, total antioxidant assays (FRAP, ABTS, DPPH), total phenolics, total flavonoids, lipid peroxidation using spectrophotometry, as well as for the heat shock protein (HSP70) using electrophoresis and silver-staining. Chilling susceptibility of lemon fruit varied with fruit source; those sourced from Ukulinga and Eston Estates were chilling resistant, while fruit from Sun Valley Estates showed chilling injury symptoms after 28 days of cold storage plus five days shelf-life. Furthermore, hot water dips (HW) 53°C, 1 μM Molybdenum (Mo) and 10 μM Mo plus HW 53°C significantly reduced chilling injury symptoms compared with the control and HW 47°C. In addition, Sun Valley Estates fruit also showed higher fruit weight loss compared with non-chilling resistant lemons. The alignment of higher fruit weight loss during storage with chilling susceptibility ascertains the use of weight loss as a non-destructive parameter for chilling susceptibility. With respect to flavedo sugars, glucose was found to be the dominant soluble sugar with multi-functional roles during cold storage. This plays a significant role in mitigating cellular stress. Chilling susceptible lemons from Sun Valley Estates had low flavedo glucose concentrations and, therefore, little conversion of glucose to ascorbic acid was possible resulting in a low antioxidant capacity. However, treatments with HW 53°C and Mo soaks seemed to enhance the enzymatic conversion of glucose to ascorbic acid leading to a higher antioxidant capacity in the flavedo of such treated fruit. Furthermore, glucose also feeds into the pentose phosphate pathway which is coupled with the shikimate pathway synthesizing secondary metabolites, especially of the phenolics group. The decrease in glucose was aligned to the levels of total phenolics, but not to that of β-carotene, naringin and hesperidin through 28 days into cold storage period. Moreover, as glucose also feeds into shikimate pathway, simultaneously an increase in proline flavedo concentration was observed. Proline is an antioxidant synthesized from glutamate; as cellular glucose decreases so does the total antioxidant capacity during cold storage. Ascorbic acid is a dominant and potent antioxidant in lemon flavedo as proven with the FRAP, ABTS and DPPH assays. Chilling resistant fruit have significantly higher ascorbic acid conversion. Furthermore, ascorbic acid also acts to generate the α-tocopheroxy radical to further important membrane-bound antioxidant, vitamin E (α-tocopherol equivalent). Furthermore, the DPPH assay was found to be effective in quantifying total antioxidants in lemon flavedo since it detects both lipophilic and hydrophilic antioxidants compared with the ABTS and FRAP assays which are bias to the estimation of liphophilic or hydrophilic antioxidants, respectively. The hot water and molybdenum treatments increased total antioxidants (DPPH assay) with reduced lipid peroxidation 7 days into cold storage and therefore, reduced chilling symptoms in fruit from Sun Valley Estates. The capacity of antioxidant to scavenge reactive oxygen species (ROS) was increased during cold storage and membrane stability significantly improved. Furthermore, putrescine as low valency polyamine was reduced as such compound acted as precursor to the synthesis of the high valency polyamines, spermine and spermidine. Chilling susceptible lemons from Sun Valley Estates showed increased soluble-conjugated polyamines as a response to stress. Furthermore, HW 53°C, 1 μM Mo and 10 μM Mo plus HW 53°C significantly increased the protein concentration and, therefore, likely also the occurrence of proteins with 70kDa (as estimator of HSP70). Additionally, the concentration of conjugated high valency polyamines was also increased, resulting in reduced chilling injury symptoms. The effect of ROS has only been viewed as damaging, while recently their role has also been viewed as stress acclamatory signalling compounds when produced concentrations below critical damaging threshold. Therefore, hot water dips seems to signals synthesis of total protein which include HSPs which then act throughout cold stress to protect other protein and channel other damaged proteins towards proteolysis. While molybdenum increased ROS production below damaging critical threshold, with ROS signalling stress acclimation by further signalling production of bioactive compound with antioxidant properties. / Thesis (Ph.D.)-University of KwaZulu-Natal, Pietermaritzburg, 2012.

The South African Citrus Industry is the second largest exporter of citrus, after Spain. The industry is under pressure to supply high quality fruit as well as to expand into new, high paying markets. However, high paying markets such as Japan and the USA require cold sterilised fruit as obligatory quarantine treatments against Mediterranean fruit fly (Ceratitis capitata) in order to reduce any possible spread of the pest. Citrus fruit originated from tropical climates and hence are chilling susceptible. Chilling injury symptoms appear as dark brown spots, pitting and/or decay when fruit are transferred to shelf temperatures; thus reducing the marketability of citrus fruit. Therefore, there is need for methods to mitigate chilling injury. Previous studies have shown silicon to mitigate many forms of stress without any hazardous effect on human health. Thus, the aim of the study was to investigate the potential of post-harvest silicon dips in mitigating chilling symptoms in citrus fruit. Briefly, fruit from two sources (Ukulinga Research Farm and Ithala Farm) were dipped in different silicon concentrations (0, 50, 150, and 250 mg ℓ-1) for 30 minutes and thereafter stored at -0.5 or 2⁰C for up to 28 days with weekly evaluation for chilling injury symptoms. Total antioxidants were determined using FRAP, ABTS, and DPPH assays under spectrophotometer. In addition, sugars, ascorbic acid, phenolics and flavonoids were analysed using High Performance Liquid Chromatography (HPLC). Fruit from Ukulinga Research Farm showed significantly higher total antioxidants (ascorbic acid total phenolics and specific flavonoids hesperidin and naringin) and sugars relative to fruit from Ithala Farm. Low concentrations of silicon dips significantly reduced the appearance of chilling injury symptoms by inducing an enzymatic conversion of glucose to ascorbic acid, thereby increasing the antioxidant capacity of chilling susceptible fruit. Moreover, silicon increased the concentration of total antioxidants, total phenolics and total flavonoids. High silicon concentrations had a negative effect on post-harvest quality of lemons by increasing fruit weight loss and electrolyte leakage, resulting in appearance of chilling symptoms. In conclusion, the study showed that silicon had potential to reduce chilling injury. However, high silicon concentrations raised concern, in particularly, on fruit appearance. / Thesis (M.Sc.Agric.)-University of KwaZulu-Natal, Pietermaritzburg, 2012.