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James R. Gorny and Adel A. Kader

The objective of this study was to compare and contrast the mode of action by which elevated carbon dioxide and/or reduced oxygen atmospheres inhibit ethylene biosynthesis. `Golden Delicious' apple fruit were placed at 0C in one of the following four atmospheres: 1) air; 2) air + 5% CO2; 3) 2% O2 + 98% N2; or 4) 2% O2 + 5% CO2 + 93% N2 and then sampled monthly for 4 months. Ethylene biosynthesis rates and in vitro ACC synthase activities were closely correlated in all treatments. In vitro ACC synthase activity and ethylene biosynthesis rates were lowest in fruit treated with 5% CO2 + 2% O2, while air-treated fruit had the highest ethylene biosynthesis rate and in vitro ACC synthase activity. Fruit treated with air + 5% CO2, or 2% O2 + 98% N2, had intermediate ethylene and in vitro ACC synthase activities. In vitro ACC oxidase was significantly different among treatments, but not as closely correlated with the ethylene biosynthesis rate as in vitro ACC synthase activity. Western blot analysis of the ACC oxidase protein was performed to determine if activity differences among treatments were correlated with the amount of enzyme present in vivo. ACC synthase and ACC oxidase mRNA transcript of abundance was determined via Northern blot analysis. Results will be discussed regarding how ethylene biosynthesis is inhibited at the molecular level by elevated CO2 and/or reduced O2.

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James R. Gorny and Adel A. Kader

Preclimacteric `Golden Delicious' apples (Malus domestica Borkh.) were stored at 0 °C in: air; air + 5% CO2; 2% O2 + 98% N2; or 2% O2 + 5% CO2 + 93% N2, and sampled monthly for 4 months to investigate the mechanism(s) by which reduced O2 and/or elevated CO2 atmospheres inhibit C2H4 biosynthesis. Ethylene biosynthesis rates and in vitro ACS activity were closely correlated in all treatments, while in vitro ACO activity significantly increased over time regardless of the treatment. Only a small amount of C2H4 biosynthesis inhibition by lowered O2 and/or elevated CO2 atmospheres could be accounted for by suppressed induction of ACO activity. Western blot analysis demonstrated that apples held for 2 months in lowered O2 and/or elevated CO2 atmospheres had significantly reduced abundance of ACO protein, compared to fruit held in air. Northern blot analysis of ACS and ACO transcript abundance revealed that reduced O2 and/or elevated CO2 atmospheres delay induction and reduce the abundance of both transcripts. Reduced O2 and/or elevated CO2 atmospheres reduce C2H4 biosynthesis by delaying and suppressing expression of ACS at the transcriptional level and by reducing the abundance of active ACO protein. Chemical names used: 1-aminocyclopropane-1-carboxylic acid (ACC), ACC synthase (ACS), ACC oxidase (ACO), ethylene (C2H4), S-adenosylmethionine (AdoMet).

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James R. Gorny and Adel A. Kader

Ethylene biosynthesis of Golden Delicious apple fruit at 20°C is rapidly inhibited by a controlled atmosphere of air + 20% CO2. However, in vitro ACC oxidase activity and ACC content were not significantly different between air and air + 20% CO2 treated fruit, To determine the in vivo effects of CO2 treatment, both in vivo and in vitro enzyme activity essays were performed in en atmosphere of air or air + 20% CO2. Western blots were also performed to quantify the amount of ACC oxidase protein present in the air and air + 20% CO2 treated fruit.

We believe that in vivo cytosolic pH changes, induced by CO2, may reduce the in vivo catalytic capacity of ACC oxidase, end hence significantly reduce ethylene biosynthesis in climacteric tissue,

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James R. Gorny and Adel A. Kader

Autocatalytic C2H4 biosynthesis in preclimacteric apple fruit (Malus domestica Borkh. `Golden Delicious') was prevented by storage in atmospheres of 20% CO2-enriched air (17% O2 + 63% N2) or 0.25% O2 (balance N2). In preclimacteric fruit, both treatments inhibited C2H2 biosynthesis by suppressing expression of ACC synthase (ACC-S) at the mRNA level. ACC oxidase (ACC-O) mRNA abundance and in vitro enzyme activity also were impaired by these treatments. However, the conversion of ACC to C2H4 never became the rate limiting step in C2H4 biosynthesis. C2H4 biosynthesis also was effectively inhibited in climacteric apple fruit kept in air + 20% CO2 or 0.25% O2. Climacteric apples also exhibited suppressed expression of ACC-S at the mRNA level, while ACC-O transcript abundance, enzyme activity, and protein abundance were reduced only slightly. ACC-S is the key regulatory enzyme of C2H4 biosynthesis and is the major site at which elevated CO2 and reduced O2 atmospheres inhibit C2H4 biosynthesis, irrespective of fruit physiological maturity. Chemical names used: 1-aminocyclopropane-1-carboxcylic acid (ACC).

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James R. Gorny and Adel A. Kader

The optimal `Bartlett' pear ripeness stage for fresh-cut processing based on flesh firmness ranges between 44.5 and 58 N (10 and 13 lbf). Use of softer pears reduces postcutting life due to flesh browning. Firmer pears may have longer postcutting life but lack good flavor. Dipping pear slices in a mixture of 2% (w/v) ascorbic acid + 1% (w/v) calcium lactate + 0.5 (w/v) cysteine (pH 7) for 5 min at 20 °C extended their shelf-life by inhibiting flesh softening and surface browning during storage at 0 °C for 10 days. After 3 days at 0 °C, ascorbic acid and cysteine residues dropped below detectable levels, while calcium content was double that of untreated slices. Preliminary sensory evaluation indicate no negative impact on flavor from this dip treatment. Exposure of intact pears to heat (35 or 40 °C) or controlled atmospheres (0.25 kPa O2 and/or 40 kPa CO2) for 24 or 48 h did not influence postcutting cut surface browning of pear slices. Storage of `Bartlett' pears at -1 °C in 2 kPa O2 (balance N2) resulted in longer postcutting life of the slices as compared to those made from air-stored pears at -1 °C. The longer the storage duration of whole pears, the shorter the shelf-life of their slices was. Fruit size did not affect the postcutting life of the pear slices, provided that they were treated with the ascorbic acid + calcium lactate + cysteine mixture. Untreated slices made from small pears exhibited surface browning faster than those made from large pears.