Analysis and evaluation of citrulline content in watermelons

Abstract: 【Objective】L-citrulline is a naturally occurring amino acid that functions in detoxification ofcatabolic ammonia and is a key element in the production of the vasodilator nitric oxide. Watermelon isconsidered to be a natural source of citrulline because of abundant citrulline in the flesh and rind. Withthe improvement of people’s living standard, functional foods have been highly valued, and watermelonvarieties with higher citrulline have been bred. Studies have been carried out to quantify the content of ci⁃trulline in different varieties with different ploidies and flesh colors, in different organs, and at fruit devel⁃opment stages. However, a problem in these studies is the lack of comparability of citrulline content because varieties evaluated were few and the studies were carried out at different growing environment in different locations. In this study, the contents of citrulline in 195 watermelon accessions planted in the samelocation were measured. According to the citrulline content level, the 195 accessions were divided into different groups, and then the distribution pattern of flesh color (white, yellow, pink, red) and watermelontypes (wild watermelons, egusi watermelons, seed watermelons, the landrace cultivars, locally bred culti⁃vars and introduced cultivars) in different groups were evaluated. Most of the traditional varieties (wild wa⁃termelons, egusi watermelons, seed watermelons, and the landrace cultivars) have been replaced by hy⁃brids and new cultivars with higher productivity and strong tolerance. Therefore, in this study, the 195 wa⁃termelon accessions were evaluated in order to recover the use of some varieties and thus increase agrobiodiversityand to screen some excellent varieties for breeding cultivars with high citrulline content.【Methods】The plants of the 195 watermelons were planted in Henan province under the same condition.The central flesh tissue from ripe fruits was used to measure the citrulline content. The tissues sampledwere immediately frozen in liquid nitrogen and quickly stored at -80 ℃ till use. Citrulline was extractedby methanol-HCl water-bathed at 55 ℃ for 30 min after the watermelon tissue had been homogenized.The extraction was added with activated carbon and then boiled for 30 min in the mixture of phosphoricacid and sulfuric acid in darkness. Coloration was developed by adding diacetyl monoxime to the mixture.A UV-visible spectrophotometer was used to determine the citrulline content in watermelon fruit indirectly.The analysis was conducted with three replicates.【Results】The results showed that the mean content ofthe 195 watermelons was 1.45 g·kg^-1, with a variation coefficient of 0.30. The cultivar‘98A13’with redflesh showed the highest citrulline concentration (2.55 g·kg^-1), while the wild watermelon‘PI 296341’with white flesh had the lowest citrulline concentration (0.49 g·kg^-1). A skewed normal distribution of citrulline content was found among the watermelon genotypes. The citrulline content in most of the varietieswas concentrated within 0.84-2.10 g·kg^-1, accounting for 82.56% of all the accessions. Cluster analysisshowed that the 195 watermelons could be divided into four categories, the very low group (I), the lowgroup (II), the high group (III), and the extremely high group (IV). Group III is the largest group containing85 (43.59%) of the watermelon genotypes tested, followed by Group II containing 52 or 26.67% of the tested genotypes. Group I contained 35 or 17.95% of the accessions, while the smallest Group IV contained 23 or 11.79% of the accessions. The citrulline content in Group IV was the highest in the 4 groups, ranging from 2.02 to 2.55 g·kg^-1, while that in Group I was the lowest, ranging from 0.49 to 0.99 g·kg^-1. The citrulline content in Group II ranged from 1.02 to 1.37 g·kg^-1 and that in Groups III from 1.39 to 1.99 g·kg^-1.The results of this study suggested that the citrulline contents in all the groups displayed a great variation.It was found that the citrulline content in white flesh watermelons was very low, especially in Group I,which contained 51.28% of all the white flesh watermelons. The yellow flesh, the pink flesh and red fleshwatermelons were mainly concentrated in Group III, accounting for 50.0%, 58.06% and 48.42% of the corresponding color types respectively. Although there was no significant difference in the citrulline contentsamong the yellow flesh, pink flesh and red flesh watermelons, the citrulline contents in brightly flesh col⁃ored watermelons (yellow, pink and red) were significantly higher than those in the white flesh ones. Thecitrulline contents in wild watermelons, egusi watermelons and seed watermelons were relatively lower,and all of them were distributed in Groups I, II and III. Wild watermelons and seed watermelons weremainly distributed in Group I, accounting for 60.0% and 50.0% of the corresponding types, respectively.The egusi watermelons were mainly in Groups I and II, which contained 40.0% of this type. However, thelandrace cultivars, bred cultivars and cultivars from abroad could be found in all of the 4 groups. The citrulline contents of them were relatively higher and all of them were in Group III, which had 44.44%,50.0% and 55.0% of the landrace cultivars, locally bred cultivars and introduced cultivars, respectively.Although there was no significant difference in citrulline content among landrace cultivars, locally bredcultivars and introduced cultivars from foreign, they were significant higher in citrulline content than wildwatermelons, egusi watermelons and seed watermelons.【Conclusion】The citrulline content exhibited largevariation among the 195 watermelon accessions, ranging from 0.49 to 2.55 g·kg^-1. According to citrullinecontent, the 195 watermelon accessions were divided into 4 groups. The citrulline contents in watermelons with colored (yellow, pink, red) flesh were significantly higher than those in white flesh ones; landrace cultivars, locally bred cultivars and introduced cultivars had higher citrulline contents than wild watermelons, egusi watermelons and seed watermelons. The citrulline contents might be related to watermelon fleshcolor and type. The difference in citrulline content among different types of watermelons is likely influenced by artificial selection and domestication of watermelons. The varieties with high citrulline contentcan be screened from locally bred cultivars, the landrace cultivars and the cultivars introduced fromabroad. From this study, 23 watermelons with high citrulline content were found valuable for breeding andgermplasm innovation.