This experiment was carried to investigate the optimum concentration of abscisic acid (ABA) treatment for enhancing fruit coloration of 'Kyoho' grapes. Cluster and berry weights showed a tendency that increased in proportion to concentration of ABA treatment, but were not significant in all treatments. Also, soluble solids and titratable acidity were not significant during fruit development in all treatments. L-phenylalanine ammonia-lyase (PAL) activity showed a tendency that decreased after rapidly increased in all treatments, and was the highest in 1000 mg/l ABA treatment at final harvest. Anthocyanin and total phenolics contents were high in proportion to ABA treatment concentration, and anthocyanin content in 1000 mg/l ABA treatment was 2.5 folds of that in control. Fructose and glucose as soluble sugars were detected, but sucrose was not detected. Both fructose and glucose contents increased during fruit development, but showed little difference in all treatments.
Hot water dipping test was conducted for chicon to restrict red discoloration of its basal part which impairs the product value during sales. Hot water dipping treatment was given to chicon for 4 min and for 8 min at $38^{\circ}C$ and for 2 min and 4 min at $42^{\circ}C$, and for 1 min and 2 min at $45^{\circ}C$, along with control (for one min at $20^{\circ}C$). The red discoloration indices of basal part of chicon during sensory evaluation on the sixth day of storage under the storage temperature at $10^{\circ}C$ was lower at $42^{\circ}C$ for 2 min, $42^{\circ}C$ for 4 min and $45^{\circ}C$ for 1 min treatments. The color change value of the basal part in chicon measured by colorimeter showed that the lowest ${\Delta}a^*$ and ${\Delta}h$ were maintained in the basal part of chicon treated at $42^{\circ}C$ for 2 min. Whereas, color changes in $42^{\circ}C$ for 2 min and $45^{\circ}C$ for 1 min treatments were significantly low as compared with that of control. The contents of total phenolic compounds which are the substances that cause red discoloration of basal part in chicon were lowest at $42^{\circ}C$ for 2 min, $42^{\circ}C$ for 4 min and $45^{\circ}C$ for 1 min treatments. The activity of phenylalanine ammonia lyase (PAL) resposible for in the synthesis of phenolic substances was the least in $42^{\circ}C$ for 2 min treatment. Whereas, PAL activity of the chicons treated a t $42^{\circ}C$ for 2 min and at $45^{\circ}C$ for 1 min were significantly lower than that of control. However, red discoloration was progressed as similar level with that of control in the basal part of chicon at $45^{\circ}C$ for 2 min. The contents of total phenolic compounds and PAL activity in this treatment were not significantly different from those in control. The polyphenol oxidase (PPO) activity which causes red discoloration of cut tissues was low in all the treatments including $42^{\circ}C$ and $45^{\circ}C$ treatment at which no inhibition effects of the red discoloration of basal part of chicon were observed. When the correlation coefficient between each investigated index was tested, most of them showed high correlation except the PPO activity and particularly and the red discoloration index and sensory evaluation ${\Delta}h$ values, and PAL activity and total phenolic compounds content were $r=0.927^{**}$, and $r=0.942^{**}$, respectively.
The effect of fungal elicitor and heavy metal salts on the production of flavonol glycosides in cell cultures of Ginkgo biloba was investigated. Among the fungi tested, Trichoderma longibrachiatum ATCC 52326 was found to be the most efficient in the production of flavonol glycosides. Kaempferol production from the elicited callus increased ten-fold as compared to the unelicited callus, while quercetin concentration of elicited cells was nine-fold higher than that of uneliceited cells in suspension cultures. The maximum quercetin concentration of 0.362㎎/l was obtained in 1.25㎎/l of the homogenate elicitor. Among the heavy metal salts tested, CuSO₄ showed a significant effect on quercetin accumulation, reaching to the concentration of 0.526 ㎎/l. Quercetin concentration increased to a maximum of l2-fold in response to CuSO₄ treatment as compared to that of untreated cells. The phenylalanine ammonia-lyase (PAL) activity and flavonol glycosides production simultaneously increased for 5 days of culture after fungal elicitor feeding, and their contents showed the same proportional patterns during the culture period. In contrast, PAL activity of cell cultures treated with CuSO₄ was almost constant during the culture period, although quercetin production increased remarkably.
Anthocyanin synthesis in strawberry (Fragaria x ananassa cv Maehyang) begins approximately 26 days postflowering and continued throughout fruit ripening. A set of cDNA clones encoding the anthocyanin biosynthetic enzymes were isolated from strawberry. A pair of primers were designed for polymerase chain reaction (PCR) through the comparison of the nucleotide sequences of homologous genes from diverse plants. Reverse transcriptase-PCRs were performed using cDNA synthesized from ripe fruit total RNA and the primers corresponding to each gene. Eight genes of the anthocyanin pathway were cloned and confirmed by sequencing to code for phenylalanine ammonia lyase (PAL), 4-cummarate CoA ligase (4CL), chalcone synthase (CHS), chalcone isomerase (CHI), flavanone-3-hydroxylase (F3H), dihydroflavonol 4-reductase (DFR), anthocyanidine synthase (ANS), UDP-glucose:flavonoid-3-O-glucosyl-transferase (UFGT). Northern analyses showed that the corresponding genes were differentially expressed during the fruit development process. All genes except PAL were predominantly expressed in fruit. Expression of PAL, DFR and ANS was detected 10 days postflowering at the early stage of fruit development, declined for a while and sharply increased 22 days postflowering then showed a peak 34 days postflowering. The other genes, however, were not expressed up to 22 or 30 days postflowering when the initial fruit ripening events occur at the time of initiation of anthocyanin accumulation. The onset of anthocyanin synthesis in ripening strawberry coincides with a coordinated induction of the anthocyanin pathway genes, suggesting the involvement of regulatory genes. We propose that at least two different regulatory mechanisms playa role in the biosynthesis of anthocyanin during color development of strawberry.
Pseudomonas fluorescens 1-94 induced systemic resistance in chickpea against Fusarium wilt of chickpea caused by Fusarium oxysporum f. sp. ciceri by the synthesis and accumulation of phenolic compounds, phenylalanine ammonia lyase(PAL) and pathogenesis related(PR) proteins(chitinase, $\beta$-1,3-glucanase and peroxidase). Time-course accumulation of these enzymes in chickpea plants inoculated with P. fluorescens was significantly(LSD, P=0.05) higher than control. Maximum activities of PR-proteins were recorded at 3 days after inoculation in all induced plants; thereafter, the activity decreased progressively. Five PR peroxidases detected in induced chickpea plants. Molecular mass of these purified peroxidases was 20, 29, 43, 66 and 97 kDa. Purified peroxidases showed antifungal activity against plant pathogenic fungi.
Riboflavin caused induction of systemic resistance in chickpea against Fusarium wilt and charcoal rot diseases. The dose effect of 0.01 to 20 mM riboflavin showed that 1.0 mM concentration was sufficient for maximum induction of resistance; higher concentration did not increase the effect. At this concentration, riboflavin neither caused cell death of the host plant nor directly affected the pathogen's growth. In time course observation, it was observed that riboflavin treated chickpea plants were inducing resistance 2 days after treatment and reached its maximum level from 5 to 7 days and then decreased. Riboflavin had no effect on salicylic acid(SA) levels in chickpea, however, riboflavin induced plants found accumulation of phenols and a greater activities of phenylalanine ammonia lyase(PAL) and pathogenesis related(PR) protein, peroxidase was observed in induced plant than the control. Riboflavin pre-treated plants challenged with the pathogens exhibited maximum activity of the peroxidases 4 days after treatment. Molecular weight of the purified peroxidase was 42 kDa. From these studies we demonstrated that riboflavin induced resistance is PR-protein mediated but is independent of salicylic acid.
Anthracnose development by Colletotrichum musae was observed to be significantly less in the fruits of the banana cultivar 'Embul' (Mysore, AAB) infected with Phyllosticta musarum than in fruits without such infections. Anthracnose disease originates from quiescent C. musae infections in the immature fruit. P. musarum incites minute, scattered spots, referred to as freckles, in the superficial tissues of immature banana peel which do not expand during maturation or ripening. P. musarum does not appear to have a direct suppressive effect on C. musae as conidia of C. musae germinate on both freckled and non-freckled fruit forming quiescent infections. Our investigations have shown that P. musarum infection induced several defence responses in fruit including the accumulation of five phytoalexins, upregulation of chitinase and ${\beta}$-1,3-glucanase, phenylalanine ammonia lyase (PAL) activity and cell wall lignification. $^1H$ and $^{13}C$ NMR spectral data of one purified phytoalexin compared closely with 4'-hydroxyanigorufone. Some of the P. musarum-induced defences that retained during ripening, restrict C. musae development at the ripe stage. This paper examines the potential of P. musarum-induced defences, in the control of anthracnose, the most destructive postharvest disease in banana.
This experiment was carried to investigate the optimum time and times of abscisic acid (ABA) treatment for the coloration in 'Kyoho' grapes. The L-phenylalanine ammonia-lyase (PAL) enzyme activities were showed highly in both ABA treatments at veraison and 10 days after veraison. However, PAL enzyme was slightly higher in ABA treatment at 10 days after veraison than in that at veraison. Anthocyanin content showed a tendency that were increased during fruit development after veraison in all treatments, and was the highest in ABA treatment at 10 days after veraison. Fructose and glucose as soluble sugars were detected by HPLC and showed little differences in all treatments. In times of ABA treatment, PAL activity showed a tendency that decreased after increased in all treatments. PAL activity in 2 times treatment of ABA was higher than other treatments. Also, anthocyanin content was highest in 2 times treatment of ABA, as 5 folds of control and folds of 1 time treatment of ABA. Both fructose and glucose contents in all ABA treatments was slightly higher than control.
When effects of exogenous hormone on hairy root cultures of Raphanus sativs cv. Chungpihongsim examined, the highest anthocyanin synthesis and disorganization were observed when 2, 4-D was supplemented to the culture medium Cytokinins showed early weak induction after transfer and ABA showed inhibitory effect and GA3 showed no effects in anthocyanin synthesis. Hormones except for 2, 4-D in 1 mg/$\ell$ concentration did not induce disorganization of hairy root and retarded growth of hairy root. Time-course changes in anthocyanin synthesis, phenylalanine ammonia-lyase activity and chalcone synthase activity were examined in culture condition contalning 2, 4-D and kinetin. In a medium containing 2, 4-D, anthocyanin synthesis began to increase on the 9th day and reaching maxima on the 18th day after transfer. Maximum peak of PAL activity appeared on the 3-9th day and another minor peak appeared on the 18th day. CHS activity increased from 9th day, reaching maximum on the 18th day and remained at a relatively high level for culture period. In a medium containing kinetin, anthocyanin synthesis increased temporarily on the 6-9th days, early days after transfer and maintained at a low level for remaining culture period. Peak of PAL activity appeared on the 6th day and CHS activity increased from the 6th days, reaching maxima about 18th day and remained at a relatively high level. In particular, addition of kinetin after preculture in hormone free medium for 2 weeks which was thought of wound healing period showed no effects in anthocyanin synthesis. This results showed that stimulation of anthocyanin synthesis by 2, 4-D and kinetin was meaningfully connected with changes of PAL, CHS activity, and then suggested rate-limiting role of CHS on anthocyanin synthesis in that there is close correlation between anthocyanin synthesis and changes of CHS activity in time-course. Besides, it is considered that cytoklnins involving kinetin stimulated anthocyanin synthesis be due to "wound response" by cutting of young roots, and that difference in time-course peak and PAL, CHS activities expressed by 2, 4-D and kinetin result from occurrence of isozyme which have different regulatory mechanism.mechanism.
Various researches on the effects of fertilization levels on functional metabolites in crop have been conducted. This review summarizes the previous studies on the relation between fertilization supply and accumulation of metabolites (phenolics, carotenoids, ascorbic acid and glucosinolates) which function as antioxidants in crop. The accumulation of phenolic compounds is related to the activation of phenylalanine ammonia lyase (PAL) in phenylpropanoid pathway. Most of the previous studies discuss that low nitrogen (N) supply activates PAL, thereby increasing the synthesis of phenolics. Similarly, high N supply leads to a decrease in ascorbic acid because of the shading effect derived from the accelerated vegetative growth under high N level. Unlike the phenolics and ascorbic acid, carotenoids are accumulated with increasing N supply. In this regard, the previous studies explain that N is a main element closely associated with formation of key enzyme for the synthesis of carotenoids. Glucosinolates are generally increased under decreasing N supply and increasing S supply. Although the previous studies show similar trends about the accumulation of metabolites by nutrient level, they also suggest that many other factors including crop types, cultivars, cultural environment (water, temperature, light, etc.) influence the accumulation of functional metabolites in crop.
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