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All parameters of the production process have been surveyed, including: cultivar, maturity stage, postharvest fruit storage; processing (malaxation time and temperature); oil storage; and finally oil storage during consumer use. Multivariate statistical analyses have been applied to identify the volatile and phenolic compounds (and other parameters) most characteristic of a certain process. It is these compounds that may be monitored more closely in any follow up studies.

Who is the report targeted at?
We see this report as forming the basis for others to further investigate the complex relationships that emerge during the production of oil from fruit, through to the finished product. It also provides useful advice to alive growers and processors on the maintenance of quality of virgin olive oil.

Aims/Objectives
This project set out to achieve the following:

1. Statistically identify cultivar differences and determine changes in volatile and phenolic profiles during fruit maturation for the production of premium quality virgin olive oil.
2. Systematically identify volatile compounds; phenolic compounds; and quality indices that significantly (p < 0.01) change with simultaneous changes in malaxation time and temperature and ultimately predict the optimum processing conditions for the transferring of the best quality attributes of the fruit to the extracted oil.
3. Determine changes in virgin olive oil quality due to different storage conditions and identify conditions that best preserve the quality and freshness of virgin olive oil.
4. Identify the critical production steps from olive fruit to oil at consumption that can be controlled to produce and maintain premium quality virgin olive oil.

Methods and results
Cultivar/maturity stage. Olive oil and fruit samples from six cultivars sampled at four different maturity stages were discriminated through statistical analysis into cultivars and maturity stages. The variables – volatile and phenolic compounds – that significantly (p< 0.01) discriminated cultivars and maturity stage groups were identified. Separation by stepwise linear discriminant analysis revealed that Manzanilla olive cultivar was separated from cultivars Leccino, Barnea, Mission, Corregiola, and Paragon, whereas cultivars Corregiola and Paragon formed a cluster. The volatile compounds hexanol, hexanal, and 1-penten-3-ol were responsible for the discrimination of cultivars. All maturity stages were discriminated, with the separation of early stages attributed to oil phenolic compounds, tyrosol and oleuropein derivatives, whereas the volatile compounds (E)-2-hexenal, hexanol, 1-penten-3-ol, and (Z)-2-penten-3-ol characterized the separation of all maturity stages and in particular the late stages. Hexanol and 1-penten-3-ol characterized the separation of both cultivars and maturity stages.

These results demonstrate that objective, instrument-based analyses are capable of measuring compounds that distinguish between cultivars and maturity stages and that oils from different fruit give different responses. This knowledge may be utilised in future studies that wish to investigate more fully the relationship between horticultural practices and sensory properties.

Post-harvest fruit storage. Frantoio olive fruits were stored at low temperature (4 ± 2°C) for 3 weeks to investigate the effect of post-harvest storage on virgin olive oil quality. Statistical analysis of variables recognized by the IOOC as measures of oil quality (FFA, PV, K232 and K270) could not explain changes in sensory quality of oils produced from stored fruit. Volatile and phenolic compounds, however, did account for observed changes in quality. Increase in concentrations of E-2- hexenal and hexanal corresponded to positive sensory quality whereas increase in E-2-hexenol and (+)-acetoxypinoresinol was associated with negative sensory quality. Volatile and phenolic compounds were also indicative of the period of low temperature fruit storage. Oleuropein and ligstroside derivatives in olive oil decreased with respect to storage time and their significant (p < 0.05) change corresponded to changes in bitterness and pungency. Z-penten-1-ol increased during low temperature fruit storage whereas 2-pentylfuran decreased.

Total volatile compounds were negatively associated with K270 and positively associated with a ketone, 6-methyl-5-hepten-2-one. These associations during low temperature storage show that olive oil quality indices were associated with volatile compounds, which in turn were associated with phenolic compounds in both the fruit and oil. The changes and associations of quality indices, sensory notes, volatile and phenolic compounds indicate that virgin olive oil quality is lost within the first week of low temperature fruit storage and re-gained at two weeks. Our research suggests that low temperature storage of olive fruit may be beneficial to the produced oil, with a possibility of increasing yield and moderating the sensory quality of olive oils. As this was a pilot study, much more work is needed to optimise storage conditions to ensure that high quality oil may be reliably produced from fruit stored prior to processing.

Malaxation time/temperature. Virgin olive oils produced at wide ranges of malaxation temperatures (15, 30, 45 and 60°C) and times (30, 60, 90 and 120 min) in a complete factorial experimental design were discriminated with stepwise linear discriminant analysis (SLDA) revealing differences in volatile and phenolic profiles with processing conditions. Virgin olive oils produced at 15°C and 60°C and malaxed for 30 min showed the most significant (p < 0.01) differences. Discrimination was based upon volatile and phenolic compounds detected in olive oils, PV, FFA, UV absorbances and oil yield.

There were different discriminating variables for processing conditions illustrating the dependence of virgin olive oil quality on malaxation time and temperature. Volatile compounds were the dominant discriminating variables. Common oxidation indicators of olive oil (PV, K232 and K270) were not among the variables that significantly (p < 0.01) changed with malaxation time and temperature.

Variables that discriminated both malaxation time and temperature were hexanal, 3,4-DHPEA-DEDA and FFA whereas 1-penten-3-ol, E-2-hexenal, octane, tyrosol and vanillic acid significantly (p < 0.01) changed with temperature only; and Z-2-penten-1-ol, (+)-acetoxypinoresinol and oil yield changed with time only. Virgin olive oil quality was significantly influenced by malaxation temperature whereas oil yield discriminated malaxation time. This study demonstrates the two modes - enzymatic and non-enzymatic - of hexanal formationduring virgin olive oil extraction. Results from this study suggest that a malaxation temperature of 30°C has benefits in terms of oil yield, while still maintaining quality. Processors may be encouraged to experiment with different malaxation times and temperatures to modify the sensory properties of their oils.

Oil storage. Virgin olive oil samples stored for 12 months in the light at ambient temperature, in the dark at ambient temperature, and at low temperature in the dark, both with and without headspace (i.e. oxygen), were separated into recognisable patterns with stepwise linear discriminant analysis (SLDA).

The discrimination with variables: volatile and phenolic compounds, free fatty acid (FFA), peroxide values (PV), K232 and K270; revealed a departure of stored oil from freshness and showed significant (p < 0.01) differences between storage conditions. Virgin olive oil stored at low temperature had characteristics closest to fresh oil while oil stored in the light showed the largest departure from freshness. Parameters that exclusively and significantly (p < 0.01) discriminated storage conditions were identified as potential markers of the storage condition. In the presence of oxygen, hexanal was a marker of storage in the light, FFA was a marker for dark storage and markers of low temperature storage were acetic acid and pentanal. In the absence of oxygen, octane was the marker for storage in the light whereas tyrosol and hexanol were markers of virgin olive oil stored in the dark, with no marker indicative of low temperature storage. E-2-hexenal, K232 and K270 were identified as markers of virgin olive oil freshness. The pronounced and rapid (< 2 months) departure from virgin status for oils stored with headspace has important implications for consumer use of oil – that once opened a bottle of oil needs to be used quickly to ensure that it remains of extra-virgin quality. A consumer education campaign may need to be devised to alert Australian olive oil users. Storage of oil in colourless glass containers may also be problematical if the oil is likely to be stored on supermarket shelves exposed to continuous visible light.

Implications
This project demonstrates the importance of the combination of objective, instrument-based analyses with statistical methods in the identification and characterisation of compounds and production steps that govern the quality and characteristics of virgin olive oil. The influence on consumed oil of important steps along the oil production process have been examined: fruit (cultivar, maturity, fruit storage); processing (malaxation time and temperature); and oil storage. Fruit cultivars have been separated based upon constituent volatile and phenolic compounds; four maturity stages have been separated by phenolic and volatile components. Storage of fruit at low temperature may be beneficial to the produced oil, with a possibility of increasing yield and moderating the sensory quality of olive oils. Processing conditions affect oil quality and yield - virgin oil quality was significantly influenced by malaxation temperature whereas malaxation time influence oil yield; results from this study suggest that a malaxation temperature of 30°C has benefits in terms of oil yield, while still maintaining quality.

The oil used by a consumer is likely no longer the oil produced at the manufacturing plant; this work indicates the sensory quality of virgin olive oil degrades upon exposure to light, and particularly degrades in a few months upon exposure to air.

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