“Sweet” or low acid pomegranates typically have been reported to have citric acid concentration less than 0.50% , but standards have not been established for low acid cultivars. Standards have been established for titratable acidity and total soluble solids of ‘Wonderful’ pomegranate. Generally, citric acid is the most abundant organic acid in pomegranate juice, so TA is expressed in citric acid equivalents. Minimum maturity of ‘Wonderful’ pomegranate fruit is 1.85% TA , so fruit are picked when fruit measure below that threshold. Maturity index is a measure of maturity in many fruit crops, including pomegranate, and is the ratio of °Brix to TA . The optimum MI for ‘Wonderful’ has been calculated to be greater than 8.1, at which point the fruit are ready for harvest. Other cultivars may have significantly different quantities of organic sugars and acids, so MI may be different for pomegranates harvested for different taste preferences and use which require fruit to be insipid, sweet, sweet-tart, tart, sour, bitter, etc. Fruit quality is not only related to sugar content, titratable acidity and spoilage, but also to phenolic compounds that lend to the fruit’s flavors, antioxidant activity, and colors . Cultivar is more influential in determining fruit juice composition than site of cultivation, year of harvest, or length of storage , so it is important to study differences among cultivars. In addition, it allows for a greater understanding of biodiversity of pomegranate germplasm. Knowing levels of phenolicsand the antioxidant activity of a cultivar’s juice is important to the beverage industry and consumers, because printed and televised commercials use antioxidant activity as the main selling point of their product. If any cultivar were to demonstrate a greater antioxidant activity than Wonderful,blueberry containers it would possibly be competitive in the pomegranate market or could be utilized by a breeder for increasing antioxidant activity. Having lower antioxidant activity than ‘Wonderful’ would make for an undesirable candidate for commercial production.
As previously mentioned, pomegranate germplasm in the United States is highly diverse and is comprised of hundreds of pomegranate cultivars sourced from domestic and international origins . Most cultivars are in the public domain with the potential to be developed into commercial cultivars. Stover and Mercure indicated that cultivars with softer seeds or lower acidity could increase consumer demand for the fruit. Wonderful, the industry standard in several countries, has relatively tart, bitter, acidic, moderately hard-seeded fruit and astringent juice compared to other cultivars previously analyzed from the USDA-ARS NCGR collection . Pomegranate cultivation in the United States is predominantly a monoculture of ‘Wonderful.’ The 14 cultivars included in the research presented herein have been described to have one or more of the following traits: unique color, soft seeds, low acidity, and/or unique flavor. Most of the cultivars in USDA germplasm have yet to be phenotyped for use by commercial growers, the food and beverage industries, and breeders. Recent developments in the California pomegranate industry have led to beverage companies predominantly desiring ‘Wonderful’ juice from growers for their products . This preference for ‘Wonderful’ juice is likely due to the clinical studies associated with this cultivar, such as the positive effect of ‘Wonderful’ juice used in conjunction with radiotherapy or surgery on men suffering from prostate cancer . Several methods exist to analyze the components and quality of fruit juices, including gas chromatography and liquid chromatography coupled with mass spectrometry, Fourier transform infrared spectroscopy , acid titration, refractometry , and spectrophotometry . All of these methods are useful, but they can be quite different in terms of sensitivity, selectivity and specificity and have disadvantages including sample destruction, incomplete sample separation in a chromatography column, and inability to profile specific quantities or classes of metabolites.
The ability of nuclear magnetic resonance to universally and quickly detect organic compounds makes it an efficacious method for the identification of differences in metabolic profile among fruit juices from different cultivars without prior separation of compounds. Additionally, NMR is advantageous because it requires relatively easy sample preparation, and allows compound identification and quantification without authentic standards . The NMR methodology has proven useful to the food and beverage industries for juice beverage quality control, including such applications as the detection of adulterants, quality control of fruit juice products during mixture analysis, and determination of juice authenticity . The pomegranate juice industry in the United States has experienced high profile lawsuits and federal investigations involving both false advertising of commercial juice composition and human health benefits from its consumption . Therefore, it is important to be able to detect beverage adulterants, such as sucrose and high fructose corn syrup, and distinguish between the juices of different fruits, vegetables and cultivars for economic, quality evaluation and public health reasons. Knowledge of a particular cultivar’s metabolic and physicochemical juice profile allows for a better understanding of pomegranate fruit juice quality, diversity of flavors and fruit maturity. For example, profiles of organic acids are known to affect flavor stability, nutrition and consumer acceptability . In regards to climate, heat has been shown to influence pomegranate fruit juice acidity. Groves planted in climates that experience hot summers typically result in fruit with reduced acidity and groves planted in climates with mild summers typically result in fruit with higher acidity . The amino acids glutamine and glutamate not only play very important physiological roles in plants, which include biochemical pathways related to amino acid synthesis, nitrogen metabolism, and ammonium detoxification, they are also important components of the human diet, with glutamine consumption becoming essential during major trauma, major surgery, sepsis, and radio- and chemotherapies .
Metabolic juice profiles might also be useful in determining fruit maturation and targeted harvest windows of pomegranate cultivars, which may be variable depending on climate and other agroecological factors. Additionally, pomegranate cultivars have differences in harvest dates, and this information is important for growers: 1) it allows them to pick fruit during harvest windows which afford the highest fruit quality for the fresh fruit, food and beverage industries; and 2) it allows growers to choose cultivars that have different maturity periods than Wonderful so they are not competing with Wonderful in domestic and international markets. Additionally, if growers start growing earlier cultivars, it could diversify the market and increase the length of the season during which high quality pomegranate fruit are available to both the food industry and consumer. The primary research objective of this dissertation was to evaluate a diverse set of preselected USDA-ARS pomegranate cultivars to determine which would meet and expand grower, consumer, and industry demands for the crop. This objective was met by: 1) conducting experiments to determine the capacity of cultivars to be propagated using conventional,best indoor plant pots inexpensive propagation methods; 2) evaluating the germplasm in field trials in hot inland and cool coastal climates to test for early orchard establishment rates, tree precocity and yield to determine commercial suitability and to aid in regional cultivar selection for growers; and 3) evaluating fruit and juice quality of the germplasm to compare important quality and nutritional traits to the industry standard ‘Wonderful.’ The pomegranate cultivars used in the following studies were: Ambrosia, Desertnyi, Eversweet, Golden Globe, Green Globe, Haku Botan, Ki Zakuro, Loffani, Nochi Shibori, Parfianka, Phoenicia, and Wonderful. All cultivars used in this experiment were located at the USDA NCGR, but the cultivars conserved in this national germplasm collection were originally sourced from different countries. ‘Ambrosia,’ ‘Eversweet,’ ‘Golden Globe,’ ‘Green Globe,’ ‘Loffani,’ ‘Phoenicia,’ and ‘Wonderful’ originated in the United States. ‘Desertnyi’ and ‘Parfianka’ originated in Turkmenistan. Haku Botan, Ki Zakuro, and Nochi Shibori originated in Japan, with Haku Botan a fruiting edible ornamental cultivar with no red pigment and Ki Zakuro and Nochi Shibori sterile cultivars, producing no fruit. All three Japanese cultivars have “double flowers” with up to hundreds of petals per flower and an upright growth habit.
Eversweet and Desertnyi have dwarf-like growth characteristics in the field, with trees typically being approximately 20% to 30% shorter or less vigorous than Wonderful as measured in the University of California, Riverside, pomegranate cultivar trials. Parfianka has a canopy with a bushy growth habit and is thornier than many other cultivars. There were no differences in bark characteristics. The first experiment included all above mentioned cultivars, and the second experiment consisted of the following cultivars: Ambrosia, Green Globe and Wonderful. Cuttings were sourced from the USDA NCGR in Winters, CA. Dormant stems were collected from basal suckers in February of two successive years for Experiments 1 and 2, respectively, placed in a plastic bag with a wet paper towel to keep them moist, driven to Davis, CA, boxed, and shipped via private delivery service from Davis, CA, to Riverside, CA, for second-day delivery. Upon arrival, the plant material was unwrapped and rew rapped with new moistened paper towels and stored at 5 to 6 °C in the dark for approximately 2 months before preparation of cuttings. All cuttings were sourced from one-year-old growth and mean stem diameter ranged from 3.8 to 5.8 mm among cultivars. Cuttings were 10.5 ± 1.0 cm long with a minimum of two nodes , treated with IBA or control, and inserted 3 to 5 cm deep in 2.5 cm x 2.5 cm plastic pots containing #4 Sunshine potting mix and perlite . Treatments were separated by block in plastic flats. Both experiments were conducted in a greenhouse at the University of California, Riverside , with mean temperatures of 24.1 °C and 26.3 °C for Experiment 1 and 2, respectively, and natural photoperiod. Cuttings were hand-watered with deionized water every day for 5 weeks and then watered as needed . Starting at week 10, the water source was switched to municipal water and all plants were treated with PlantexTM water soluble fertilizer, which contained N-P-K plus micronutrients, at a rate of 0.2 g·L-1 nitrogen. This experiment was conducted to determine if differences in rooting percentages and vegetative growth attributes exist among cultivars. The experiment ran from 11 March until 09 August of year 1 and consisted of a randomized complete block design with eight blocks. Each block included four hardwood stem cuttings of each cultivar, totaling 32 cuttings per cultivar. All cuttings were treated for 5 s to a depth of 2 to 4 cm in a gelatinous formulation of 3 g×L -1 IBA before insertion in the rooting medium. This experiment was conducted to test effects of IBA concentration on rooting percentages and vegetative growth attributes of Wonderful , Green Globe, and Ambrosia . Experiment 2 ran from 02 April until 28 December 2014 and used a RCBD with four blocks, each with four cuttings per cultivarIBA combination, totaling 48 cuttings for each of the three cultivars. IBA was applied in the same manner as in experiment 1, except that cuttings were dipped in either DI water or IBA at 1.5 g×L -1 or 3 g×L -1 IBA before insertion in potting mix. Attributes measured in both experiments included initial stem diameter, rooting percentage, plant height, number of shoots, and total shoot growth . Rooting percentages were based on cuttings that successfully rooted and developed into plants. Plant height, number of shoots, and total shoot growth were assessed at the end of each experiment: day 154 for Experiment 1 and day 270 for Experiment 2. Experiment 2 was conducted over a longer time to allow for assessment of growth after a full growing season. Plant height was measured using the top of the potting container as the reference point to determine if there were differences among cultivar phenotypes for upright versus drooping growth habit. Wonderful pomegranate grows in willowy tree form, with a tall, often thin, spreading and bending branching pattern but there are cultivars with upright growth habit, so this growth habit trait was assessed by measuring both apical shoot growth and plant height. Number of primary shoots represented a count of shoots growing from above ground, lateral buds on the cutting. Attributes measured at the end of Experiment 1 were apical shoot growth , branching , and relative chlorophyll content . SPAD values were used to assess plant health by quantifying leaf relative chlorophyll content using a SPAD-502 chlorophyll meter . Attributes measured at the end of Experiment 2 were total leaf area and root dry mass. Total leaf area was measured with a LI-COR Model 3100 Area Meter . Dry root mass was determined by removing the roots from the plant, drying the roots in a forced air oven to constant weight, and weighing the dried roots.