Banana sap has some special properties relating to various phenomena such as browning of fruits after harvesting, permanent staining of cloth and fibers, and antioxidant and antibleeding properties. Analysis of banana sap using high-performance liquid chromatography-electrospray ionization-mass spectrometry (HPLC-ESI-MS) indicated the presence of phenolic and aromatic amino compounds of interest due to their special properties. With the online positive electrospray ionization mode (ESI), the possible structures of specific compounds were determined from the fragmentation patterns of each particular ion appearing in the mass spectra. The major compounds revealed from the sap of banana accessions, namely, Musa balbisiana , Musa laterita , Musa ornata , and Musa acuminata , and some cultivars were apigenin glycosides, myricetin glycoside, myricetin-3-O-rutinoside, naringenin glycosides, kaempferol-3-O-rutinoside, quercetin-3-O-rutinoside, dopamine, and N-acetylserotonin. The results indicated that there was a variety of phenolic and aromatic amino contents in many banana species. These compounds were reported to relate with biological activities. Moreover, the identities of these phytochemical compositions may be used as markers for banana diet, the assessment of physiochemical status, or the classification of banana clones.

Anthocyanins were isolated from male bracts of 10 wild species of bananas (Musa spp. and Ensete spp.) distributed in Thailand. Six major anthocyanin pigments were identified by high performance liquid chromatography (HPLC), mass spectrometry (MS), and tandem mass spectrometry (MS/MS). They are delphinidin-3-rutinoside (m/z 611.2), cyanidin-3-rutinoside (m/z 595.8), petunidin-3-rutinoside (m/z 624.9), pelargonidin-3-rutinoside (m/z 579.4), peonidin-3-rutinoside (m/z 608.7), and malvidin-3-rutinoside (m/z 638.8). On the basis of the types of pigment present, the wild bananas can be divided into 5 groups. The first group comprises M. itinerans, Musa sp. one, Musa sp. two, and M. acuminata accessions, which contain almost or all anthocyanin pigments except for pelargonidin-3-rutinoside, including both nonmethylated and methylated anthocyanins. The second group, M. acuminata subsp. truncata, contains only malvidin-3-rutinoside while the third group, M. coccinea, contains cyanidin-3-rutinoside and pelargonidin-3-rutinoside. The forth group, M. acuminata yellow bract and E. glaucum do not appear to contain any anthocyanin pigment. The fifth group consists of M. balbisiana, M. velutina, M. laterita, and E. superbum which contain only nonmethylated anthocyanin, delphinidin-3-rutinoside, and cyanidin-3-rutinoside. Total anthocyanin content in the analyzed bracts ranged from 0-119.70 mg/100 g bract fresh weight. The differences in the type of anthocyanin and variation in the amounts present indicate that wild bananas show biochemical diversity, which may be useful for identifying specific groups of bananas or for clarifying the evolution of flavonoid metabolism in each banana group.

Bananas in Thailand have been surveyed by our team to be at least 140 cultivars in the plantations, 10 wild species and, 4 introduced species. To characterize the genetic relationship of species and cultivars, a set of novel SSR markers was developed. Totaling 53 clones containing SSR motifs were isolated from SSR-enriched library of wild Musa balbisiana Colla 'Tani' (BB). Selected positive clones were used to design 28 primer pairs for amplification of 12 wild and 82 cultivar accessions with genome designations AA, AB, AAA, AAB, ABB, and BBB. These SSR markers loci were homology searched to the banana genomes to map their locations. The seven-sets multiplex PCR approach using four fluorescent-labeled universal primers were utilized for cost effectiveness. Capillary fragment analysis yielded the accurate size of amplicons for evaluation of particular patterns for each cultivar. Phylogram and Structure analysis presented the specific genotype of genome groups (A and B genotypes, polyploid hybrid genomes) and cultivar groups. By A:B specific alleles ratio, accurate genome designations of hybrids can be determined. Additionally, a marker, characterized to be partial plastid ycf2 gene, indicated the maternal identification of hybrid cultivars. One SSR marker was also preliminary tested with some wild species and advised to be the candidate fingerprinting marker for species identification. In conclusion, SSR marker sets developed here proved their exploitation in detailed identity and relationship of cultivated bananas, which would be useful for genetic conservation and ongoing breeding programs in Thailand and other areas.

An enzymatic method for specific determination of stevioside content was established. Recombinant β-glucosidase BT_3567 (rBT_3567) from Bacteroides thetaiotaomicron HB-13 exhibited selective hydrolysis of stevioside at β-1,2-glycosidic bond to yield rubusoside and glucose. Coupling of this enzyme with glucose oxidase and peroxidase allowed for quantitation of stevioside content in Stevia samples by using a colorimetric-based approach. The series of reactions for stevioside determination can be completed within 1 h at 37 °C. Stevioside determination using the enzymatic assay strongly correlated with results obtained from HPLC quantitation (r = 0.9629, n = 16). The percentages of coefficient variation (CV) of within day (n = 12) and between days (n = 12) assays were lower than 5%, and accuracy ranges were 95-105%. This analysis demonstrates that the enzymatic method developed in this study is specific, easy to perform, accurate, and yields reproducible results.

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