Plant material

Three sets of experiments were conducted as summarised in Table 5.1. In experiments 1 and 3, ‘Owairaka Red’ roots were sourced from Delta Produce, Dargaville, New Zealand. In experiment 2, roots of ‘Clone 1820’ were used. ‘Clone 1820’ was selected because it contains substantial amounts of phenolic acids, carotenoids, and anthocyanin. The ‘Clone 1820’ roots were sourced from Plant and Food Research, Pukekohe, New Zealand. All cultivars were harvested in April 2013. Roots were transported to Massey University, Palmerston North, within 24 h of harvest. Roots without visible defects were cured at 30 °C and 90 - 95% relative humidity, for four days. ‘Owairaka Red’ roots were stored for 4 weeks and sampling was conducted every week as repeated measures. ‘Clone 1820’ roots were stored for a longer period (8 weeks) to allow an extended period in which to monitor any phytochemical changes due to exposure to ethylene and 1-MCP treatments. All the measures for ‘Clone 1820’ were non-repeated. Sensory evaluations (experiment 3) were undertaken using ‘Owairaka Red’ roots. By the time of the experiment, the roots had been in store at 14 - 15 °C for about 7 months.

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Table 5.1: Sweetpotato cultivars used and parameters measured in the three experiments

Parameters Exp.1 Exp.2 Exp.3

Weight loss ݱ ݱ -

Sprout numbers & length ݱ ݱ -

Respiration rate _{ݱ ݱ} - Rot incidence _{ݱ ݱ} - Internal colour ݱ ݱ - Sugars ݱ ݱ - Carotenoids - ݱ - Phenolic acids ݱ ݱ - Anthocyanin - ݱ - Sensory test - - ݱ

Cultivar Owairaka Red Clone 1820 Owairaka Red

Storage duration 4 weeks 8 weeks 4 weeks

5.2.2

1-MCP treatment

1-MCP was applied according to Agro Fresh guidelines; in brief, 86.9 mg of 0.14% 1- MCP powder (SmartfreshTM, Agro fresh Inc., USA) was placed in an airtight 100 mL syringe fitted with a 3-way Luer stopcock. Warm milliQ water (2 mL) was injected into the syringe to dissolve the powder. Following vigorous shaking (2 min), all the contents of the syringe were injected into closed high-density polyethylene barrels, each containing 5 kg of sweetpotato roots. Control samples were treated in the same manner, excluding the application of 1-MCP. Following 24 h of incubation at 20 ˚C, the barrels were vented outdoors. Thereafter, the roots were divided into the designated treatments, and stored in a flow-through system, with either continuous air or ethylene (10 µL L-1), for 4 and 8 weeks using ‘Owairaka Red’ and ‘Clone 1820’ respectively. Two green- breaker tomatoes were included in each of the 1-MCP treatment barrels as positive biological indicators of 1-MCP responses. The treated and untreated tomatoes were stored at room temperature (20 ˚C), and their ripening was monitored for 3 weeks. Ripening of 1-MCP treated tomatoes was significantly delayed (Figure 5.1) indicating the efficacy of applied 1-MCP.

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Figure 5.1: Visual appearance of tomatoes before treatment (+/- 1-MCP) and after 2 weeks storage at room temperature (20 °C).

5.2.3

Ethylene application

Ethylene (10 µL L-1) was applied continuously at a constant flow rate of 720 mL min-1 into the 56 L airtight barrels in a flow-through system, for the entire storage period. The desired concentration of ethylene was obtained by mixing 5000 µL L-1 ethylene (in 20.7% oxygen in nitrogen gas) (BOC Ltd, Auckland, New Zealand) and compressed air using a purpose built gas mixer with control needle valves and a pressure gauge to regulate the flow rate. The gas mix was passed through an 1800 mL

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jar containing 750 mL of 43% glycerol in water to humidify the gas to around 85 % RH before continuing to each barrel. The concentrations of the mixed gases were measured on a weekly basis using a gas chromatograph (GC) (Shimadzu GC-2014, Shimadzu Scientific Instruments, Auckland, New Zealand). The GC was calibrated using a 10.1 ± 0.5 µL L-1 ethylene standard (BOC Ltd., Auckland, New Zealand). The data were acquired using Shimadzu GC Solutions software.

5.2.4

Respiration measurement

Respiration was measured as the amount of carbon dioxide accumulated over time. Sweetpotato roots (3 roots/replicate/treatment) were placed in 1800 mL glass jars with an airtight lid fitted with a rubber septum. Using a 1 mL syringe, gas was withdrawn from the headspace just after sealing the jars and again after 30 min. Carbon dioxide concentration was determined using an infra-red CO₂ transducer (Analytical Development Company, Hoddesdon, UK), with O₂-free Nitrogen as a carrier gas (flow rate 35 mL min-1). The output signal was analysed by an integrator (Hewlett Packard, Model 3394A). Respiration rates were calculated taking into account accumulation time, sample weight, and the free volume of the jar. The final respiration rates were a mean of three replicates, each replicate consisting of three roots. The equipment was calibrated with β-standard 0.49 ± 0.01% CO₂ (BOC Ltd, Auckland, New Zealand).

5.2.5

Weight loss

Weight loss (%) during storage was measured as previously described in section 2.3.1.

5.2.6

Sprout growth

Sprout number per root and length (mm) was assessed as previously described in section 2.3.2.

5.2.7

Rot incidence

Roots with visible signs of rots were counted as rotten and expressed as a percentage incidence relative to the initial root number.

5.2.8

Colour measurement

At each sampling time, six roots were randomly selected from each treatment and halved longitudinally. One half was used for raw colour measurement, and the other half was cooked for 10 min (total quantity of sweetpotato in the oven 550 to 600g) using a domestic microwave (850 W Panasonic Model NN-7852, Matsushita

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Electrical Industrial co., Japan) and allowed to cool before measurement. Colour measurements were performed using a reflectance spectrophotometer (Model CM- 2600D, Konica Minolta Sensing Inc., Japan) equipped with an 8 mm diameter measuring head and D65 as the light source. The spectrophotometer was calibrated with a standard white colour surface. Three measurements were taken from each piece of root and averaged to give one reading per root.

5.2.9

Phytochemical analysis

The phytochemicals measured were carotenoids, phenolic acids, and anthocyanins. Samples were prepared and analysed as described previously in section 2.4.