Active recombinant thioredoxin h protein with antioxidant activities from sweet potato (Ipomoea batatas [L.] Lam Tainong 57) storage roots

Recombinant thioredoxin h (Trx2) overproduced in Escherichia coli (M15) was purified by Ni2+-chelated affinity chromatography. The molecular mass of Trx2 is approximately 1.4 kDa as determined by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. Total antioxidant status, 1,1-diphenyl-2-picrylhydrazyl (DPPH) staining, reducing power method, Fe2+-chelating ability, ferric thiocyanate (FTC) method, and protection of calf thymus DNA against hydroxyl radical-induced damage were studied. The thioredoxin h protein with a concentration of 12.5 mg/mL exhibited the highest activity (expressed as 0.37 +/- 0.012 mM ABTS* radical cation being cleared) in a total antioxidant status test. In the DPPH staining thioredoxin h appeared as white spots when it was diluted to 50 mg/mL (a final amount of 15 microg). Like the total antioxidant status, the reducing power, Fe2+-chelating ability, FTC activity, and protection against hydroxyl radical-induced calf thymus DNA damage were found with the thioredoxin h protein. It was suggested that thioredoxin h might contribute to its antioxidant activities against hydroxyl and peroxyl radicals.     

Antioxidant activities of trypsin inhibitor, a 33 KDa root storage protein of sweet potato (Ipomoea batatas (L.) Lam cv. Tainong 57).

Trypsin inhibitors (TIs), root storage proteins, were purified from sweet potato (Ipomoea batatas[L.] Lam cv. Tainong 57) roots by trypsin affinity column according to the methods of Hou and Lin (Plant Sci. 1997, 126, 11-19 and Plant Sci. 1997, 128, 151-158). A single band of 33 kDa TI was obtained by preparative sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) gels. This purified 33 kDa TI had scavenging activity against 1,1-diphenyl-2-picrylhydrazyl (DPPH) radical. There was positive correlation between scavenging effects against DPPH (2 to 22%) and amounts of 33 kDa TI (1.92 to 46 pmol). The scavenging activities of 33 kDa TI against DPPH were calculated from linear regression to be about one-third of those of glutathione between 5 and 80 pmol. Using electron paramagnetic resonance (EPR) spectrometry for hydroxyl radical detection, it was found that 33 kDa TI could capture hydroxyl radical, and the intensities of EPR signal were significantly decreased from 1.5 to 6 pmol of 33 kDa TI compared to those of the controls. It is suggested that 33 kDa TI, one of the sweet potato root storage proteins, may play a role as an antioxidant in roots and may be beneficial to health when it is consumed.     

Assessing genetic diversity of sweet potato (Ipomoea batatas (L.) Lam.) cultivars from tropical America using AFLP

The sweet potato genebank at the International Potato Center (CIP) maintains 5,526 cultivated I. batatas accessions from 57 countries. Knowledge of the genetic structure in this collection is essential for rational germplasm conservation and utilization. Sixty-nine sweet potato cultivars from 4 geographical regions (including 13 countries) of Latin America were randomly sampled and fingerprinted using AFLP markers. A total of 210 polymorphic and clearly scorable fragments were generated. A geographic pattern of diversity distribution was revealed by mean similarity, multidimensional scaling (MDS), and analysis of molecular variance (AMOVA). The highest genetic diversity was found in Central America, whereas the lowest was in Peru-Ecuador. The within-region variation was the major source of molecular variance. The between-regions variation, although it only explains 10.0% of the total diversity, is statistically significant. Cultivars from Peru-Ecuador, with the lowest level of within region diversity, made the most significant contribution to the between region differentiation. These results support the hypothesis that Central America is the primary center of diversity and most likely the center of origin of sweet potato. Peru-Ecuador should be considered as a secondary center of sweet potato diversity.     

An arsenate reductase homologue possessing phosphatase activity from sweet potato (Ipomoea batatas [L.] Lam): kinetic studies and characterization

A cDNA encoding a putative arsenate reductase homologue (IbArsR) was cloned from sweet potato (Ib). The deduced protein showed a high level of sequence homology (16-66%) with ArsRs from other organisms. A 3-D homology structure was created based on AtArsR (PDB code 1T3K ) from Arabidopsis thaliana. The putative active site of protein tyrosine phosphatase (HC(X)(5)R) is conserved in all reported ArsRs. IbArsR was overexpressed and purified. The monomeric nature of the enzyme was confirmed by 15% SDS-PAGE and molecular mass determination of the native enzyme via ESI Q-TOF. The IbArsR lacks arsenate reductase activity but possesses phosphatase activity. The Michaelis constant (K(M)) value for p-nitrophenyl phosphate (pNPP) was 11.11 mM. The phosphatase activity was inhibited by 0.5 mM sodium arsenate [As(V)]. The protein's half-life of deactivation at 25 °C was 6.1 min, and its inactivation rate constant K(d) was 1.1 × 10(-1) min(-1). The enzyme was active in a broad pH range from 4.0 to 11.0 with optimum activity at pH 10.0. Phosphatase would remove phosphate group from nucleic acid or dephosphorylation of other enzymes as regulation signaling.     

Inhibition of reactive nitrogen species in vitro and ex vivo by trypsin inhibitor from sweet potato 'Tainong 57' storage roots

Peroxynitrite (ONOO-), formed from a reaction of superoxide and nitric oxide, is one of the most potent cytotoxic species known to oxidize cellular constituents including essential proteins, lipids, and DNA. ONOO- induces cellular and tissue injury, resulting in several human diseases such as Alzheimer's disease, atherosclerosis, and stroke. Due to the lack of endogenous enzymes responsible for ONOO- scavenging activity, finding a specific ONOO- scavenger is of considerable importance. In this study, the ability of trypsin inhibitor (TI), isolated from sweet potato storage roots (SPTI), to scavenge *ON and ONOO- was investigated. The data obtained show that TI generated a dose-dependent inhibition on production of nitrite and superoxide radicals. The IC50 value of TI on superoxide radical was 143.2 +/- 4.29 microg/mL. SOD activity staining was used to confirm SOD activity of SPTI. SPTI also caused a dose-dependent inhibition of the oxidation of dihydrorhodamine 123 (DHR) by peroxynitrite. A calculated IC50 value of 809.1 +/- 32.36 microg/mL was obtained on the inhibition of peroxynitrite radical. Spectrophotometric analyses revealed that TI suppressed the formation of ONOO--mediated tyrosine nitration through an electron donation mechanism. In further studies, TI also showed a significant ability to inhibit nitration of bovine serum albumin (BSA) in a dose-dependent manner. In vivo TI inhibited lipopolysaccharide-induced nitrite production in macrophages in a concentration-dependent manner with an IC50 value of 932.8 +/- 29.85 microg/mL. The present study suggested that TI had an efficient reactive nitrogen species scavenging ability. TI might be a potential effective NO and ONOO- scavenger useful for the prevention of NO- and ONOO--involved diseases.     

Potential chemopreventive properties of extract from baked sweet potato (Ipomoea batatas Lam. Cv. Koganesengan)

The extract from baked sweet potato (Ipomoea batatas Lam. cv. Koganesengan) showed potential cancer-preventing effects. The extract was partially fractionated to four fractions (I, II-a II-b, and III) by Sephadex G-25 gel chromatography. The cytotoxicity against human myelocytic leukemia HL-60 cells, the suppression of TPA-induced transformation in mouse skin JB6 C141 cells, the apoptosis inducing activity in HL-60 cells, and the scavenging capacity against DPPH radical were tested on the four fractions. Fractions II-a and III showed markedly strong radical scavenging effects on the DPPH radical, coinciding with the high content of total phenolic compounds in the fractions. Both of these fractions suppressed strongly the proliferation of HL-60 cells with apoptosis induction in a dose-dependent manner. Moreover, the two fractions markedly blocked TPA-induced cell transformation in the JB6 cell line. Taken together, these data suggest that the water extract from baked sweet potato had potential chemopreventive properties.     

Changes in polyphenolic content and radical-scavenging activity of sweet potato (Ipomoea batatas L.) during storage at optimal and low temperatures

Polyphenolic content and radical-scavenging activities (RSA) of four sweet potato (Ipomoea batatas L.) cultivars were characterized after storage at optimal (15 degrees C) or low temperature (5 degrees C) for 0, 13, 26, and 37 days. The polyphenolic content increased during storage in three cultivars but not in 'Murasakimasari'. The change in 1,1-diphenyl-2-picrylhydrazyl radical-scavenging activity (DPPH-RSA) correlated very well with polyphenolic content. The increases in polyphenolics and the RSA in 'Benimasari' were significantly greater during storage at 5 degrees C than at 15 degrees C. The main polyphenolic components in all cultivars were chlorogenic acid (ChA) and 3,5-di-O-caffeoylquinic acid (3,5-diCQA). ChA level increased more at 5 degrees C than at 15 degrees C, whereas that of 3,5-diCQA was greater at 15 degrees C. Caffeoylquinic acids and RSA in 'Murasakimasari', which contains a large amount of anthocyanin in flesh tissue, were extremely high at the beginning of storage and remained nearly constant or decreased over time. A non-caffeoylquinic acid component that increased during storage, especially in 'J-Red' at 15 degrees C, was purified by successive chromatographic steps. The isolate was identified as caffeoyl sucrose [CSu, 6-O-caffeoyl-(beta- d-fructofuranosyl-(2-->1))-alpha-D-glucopyranoside] by fast atom bombardment-mass spectroscopy (FAB-MS), infrared spectroscopy (IR), and nuclear magnetic resonance spectroscopy (NMR). These results suggest that storage under cultivar-dependent, controlled temperature is one approach for increasing desirable physiologic function associated with RSA of polyphenolic compounds in sweet potato roots.     

Structural characterization and hypoglycemic effects of arabinogalactan-protein from the tuberous cortex of the white-skinned sweet potato (Ipomoea batatas L.)

An arabinogalactan-protein (WSSP-AGP) was isolated from the tuberous cortex of the white-skinned sweet potato (WSSP; Ipomoea batatas L.). It consists of 95% (w/w) carbohydrate and 5% (w/w) protein with high contents of hydroxyproline, alanine, and serine. Its sugar composition is α-L-Rha:α-L-Ara:β-D-Gal:β-D-GlcA in a molar ratio of 1.0:4.1:7.6:1.3. Its weight-average molecular weight was estimated to be 126,800 g/mol by high-performance size exclusion chromatography coupled with multiangle laser light scattering. Structural analysis indicated that WSSP-AGP is a (1→3)-β-D-galactan highly branched at O-6 with (1→6)-β-D-galactan, in which the branched chains are substituted at the O-3 position with α-L-Araf-(1→ and α-L-Araf-(1→5)-α-L-Araf-(1→ and at the O-6 position typically with α-L-Rhap-(1→4)-β-D-GlcAp-(1→ as terminating groups. Continuous administration of WSSP-AGP to KKAy mice significantly lowered fasting plasma glucose levels. This indicates that WSSP-AGP plays an important role in the hypoglycemic effects of WSSP.     

Sweet potato [Ipomoea batatas (L.) Lam.] cultivated as tuber or leafy vegetable supplier as affected by elevated tropospheric ozone

Sweet potato cultivars respond differently to elevated tropospheric ozone concentrations of ca. 130 mug m (-3), 8 h a day for 4 weeks, which affects their selection for cultivation. In the first cultivar presented here, an adequate leafy vegetable supplier, the ozone load resulted in a shift of biomass to maintain the canopy at the expense of tuber development. Starch content of leaves was reduced, indicating an impairment of quality, but carotenoid content remained stable. The second cultivar may be grown for tuber production. Although the ratio tuber/plant remained stable under ozone, tuber yield and its starch content were significantly reduced. The lower starch content indicated a worse quality for certain industrial processing, but it is desirable for chip production. Elevated tropospheric ozone concentrations also influenced free amino acids and macronutrient contents of tubers, but these modifications were of minor significance for tuber quality in the second cultivar.     

Analysis of genetic diversity in a sweet potato (Ipomoea batatas L.) germplasm collection from Tanzania as revealed by AFLP

Sweet potato (Ipomoea batatas L.) is the fifth most important crop in the developing countries after rice, wheat, maize and cassava. The amplified fragment length polymorphism (AFLP) method was used to study the genetic diversity and relationships of sweet potato accessions in the germplasm collection of Sokoine University of Agriculture, Morogoro and Sugarcane Research Institute, Kibaha, Tanzania. AFLP analysis of 97 sweet potato accessions using ten primer combinations gave a total of 202 clear polymorphic bands. Each one of the 97 sweet potato accessions could be distinguished based on these primer combinations. Estimates of genetic similarities were obtained by the Dice coefficient, and a final dendrogram was constructed with the un-weight pair-group method using arithmetic average. AFLP-based genetic similarity varied from 0.388 to 0.941, with a mean of 0.709. Cluster analysis using genetic similarity divided the accessions into two main groups suggesting that there are genetic relationships among the accessions. Principal Coordinate analysis confirmed the pattern of the cluster analysis. Analysis of molecular variance revealed greater variation within regions (96.19%) than among regions (3.81%). The results from the AFLP analysis revealed a relatively low genetic diversity among the germplasm accessions and the genetic distances between regions were low. A maximally diverse subset of 13 accessions capturing 97% of the molecular markers diversity was identified. We were able to detect duplicates accessions in the germplasm collection using the highly polymorphic markers obtained by AFLP, which were found to be an efficient tool to characterize the genetic diversity and relationships of sweet potato accessions in the germplasm collection in Tanzania.     

Calf thymus DNA-binding ability study of anthocyanins from purple sweet potatoes ( Ipomoea batatas L.)

A total of 10 anthocyanin compounds were identified from five purple sweet potato ( Ipomoea batatas L.) varieties, Qunzi, Zishu038, Ji18, Jingshu6, and Ziluolan, by high-performance liquid chromatography-tandem mass spectrometry (HPLC-MS/MS) to assess their calf thymus DNA-binding ability in vitro. The interaction between anthocyanins and calf thymus DNA in Tris-HCl buffer solution (pH 6.9) was evaluated by fluorescence spectroscopy. Using ethidium bromide (EB) as a fluorescence probe, fluorescence quenching of the emission peak was seen in the DNA-EB system when anthocyanins were added, indicating that the anthocyanins bound with DNA. The acylated groups influenced the ability of the interaction with DNA. Anthocyanins from purple sweet potato with more acylated groups in sorphorose have a stronger binding ability with DNA.     

甘薯抗茎线虫病基因的AFLP标记的开发

以甘薯抗茎线虫病品种徐781和感茎线虫病品种徐薯18的杂交F1分离群体的186株单株为材料,利用分离群体分组分析法（BSA法）和AFLP技术,在抗感池中共筛选了800对AFLP引物,结果表明其中245对引物具有多态性。用这245对引物检测两亲本以及建池单株,发现引物组合E2M23和E33M20分别在抗病单株中扩增出1条在感病单株中未出现的特异条带,长度分别约为500 bp和200 bp,认为这2个AFLP标记与甘薯抗茎线虫病基因连锁,分别命名为E2M23500和E33M20200。根据这2个AFLP标记对F1代186个单株的扩增结果,经Mapmaker 3.0软件分析,发现这2个分子标记与抗茎线虫病基因位于同一连锁群并紧密连锁,它们与抗茎线虫病基因间的遗传距离分别为6.94 cM和11.1 cM。用这2个分子标记对10个中国甘薯主栽品种进行检测,所得结果与常规方法鉴定结果完全一致,表明2个分子标记可用于甘薯抗茎线虫病分子标记辅助育种。     

Oxidative comparison of emulsion systems from fish oil-based structured lipid versus physically blended lipid with purple-fleshed sweet potato (Ipomoea batatas L.) extracts

The effects of the purple-fleshed sweet potato extract (PFSPE) on oxidation stabilities of a model oil-in-water emulsion prepared with enzymatically synthesized fish oil-soybean oil structured lipid (SL) versus physically blended lipid (PBL) without modification were evaluated. The anthocyanins in PFSPE were analyzed and identified by HPLC-MS. The fatty acid composition of SL was similar to that of PBL, except palmitic acid (1.48 in PBL and 9.61% in SL) and linoleic acid (62.47 in PBL and 49.58% in SL). Peonidin 3-caffeoylsophoroside-5-glucoside, peonidin-3-(6',6'-caffeoylferuloylsophoroside)-5-glucoside, peonidin-dicaffeoylsophoroside-5-glucoside, peonidin 3-(6',6"-caffeoyl-p-hydroxybenzoylsophoroside)-5-glucoside were identified as the major anthocyanin compounds in PFSPE. Different levels (200, 500, 1000 ppm) of PFSPE were added into both SL- and PBL-based emulsions, with 200 ppm catechin as comparison. Oxidation was monitored by measuring the peroxide value and thiobarbituric acid reactive substances. The antioxidant activity of PFSPE increased with an increased concentration, the concentration of 1000 ppm showed high antioxidant ability similar to that of catechin in both PBL- and SL-based oil-in-water emulsions. It is notable that the SL-based emulsion appeared to have better oxidative stability than the PBL-based emulsion.     

Differences in mycorrhizal infection and P uptake of sweet potato cultivars (Ipomoea batatas L.) during their early growth in three soils

Summary Mycorrhizal infection in the roots of 10 sweet potato cultivars was assessed 7 weeks after planting in three soils collected from Ibadan, Fashola and Onne in southern Nigeria, three soils which contained 21.0, 7.8 and 54.8 mg P kg^–1, respectively. Mycorrhizal infection averaged 17% in the soil from Ibadan, 24% in the soil from Fashola and 7% in the acid soil from Onne. The plants grown in the Fashola soil contained the same percentage of P as plants grown in the Onne soil. Although the percentage of P in sweet potato was lowest in the Ibadan soil, shoot dry weights were 35% higher in this soil than in the other two soils. There was no correlation between the level of mycorrhizal infection and plant dry weight in the partially sterilized soil from Ibadan. Sweet potato inoculated in this soil with infected roots of Leucaena leucocephala showed a higher level of mycorrhizal infection than uninoculated plants. Dry-matter production was, however, the same for all treatments. The sweet potato cultivars differed in their level of mycorrhizal infection and in their response to applied P. Cultivars TIS 2498 and TIS 70357 consistently showed the lowest percentage of infection; and TIb 4, TIS 8441 and TIS 8524 showed infection levels above 20% in the Fashola and Ibadan soils. When the low-yielding cultivar, TIb 4, and an improved clone, TIS 9265, were grown in the presence of 50 and 100 mg single superphosphate per kg soil, TIb 4 produced more dry matter in the presence of P fertilizer than it did without the fertilizer. Growth and mycorrhizal infection of TIS 9265 were not affected by the fertilizer.     

Growth and acquisition of Na,K and Ca in some elite sweetpotato [Ipomoea batatas (Lam.) L.] genotypes under salinity stress

Soil salinity is a concern in the wake of climate change challenges due to rising sea levels and coastal salinity in Papua New Guinea. A greenhouse experiment was conducted in Split Plot design, with five elite sweet potato genotypes (main-plot factors) and three levels of sodium chlroide (NaCl) concentrations (sub-plot factors) replicated six times. The vine cuttings of genotype RAB 45 showed very low mortality percentage (33%) at 600 mM NaCl concentration. At salinity level of 200 mM NaCl, aerial dry biomass of the genotypes was inversely but significantly (r = –0.40; p < 0.05) related to the accumulation of sodium (Na⁺) in the tissues. The Na⁺ accumulation in the tissues was antagonistic to the potassium (K⁺) and calcium (Ca²⁺) ions. Among the sweetpotato genotypes, Na⁺/K⁺ ratio decreased in the following order: RAB 45> KAV 11 > Northern Star > DOY 2 > L 46, which was more or less corroborated with the trend in the aerial dry matter.     

Genetic diversity in sweetpotato [Ipomoea batatas (L.) Lam.] in relationship to geographic sources as assessed with RAPD markers

Available evidence shows that sweetpotato originated from either Central or South American lowlands with subsequent dispersal to North America, Europe, Africa, Asia and the pacific islands. A total of 71 polymorphic RAPD molecular markers were used to assess the genetic relationships amongst 74 sweetpotato varieties originating from a total of 23 sweetpotato producing countries within six geographical regions, namely, South America, Central America/Caribbean, United States of America (USA), East Africa, Asia and Oceania. An Analysis of Molecular Variance (AMOVA) indicated that 93.4% of the total variance was due to the differences between genotypes within regions. The difference between regions was significant (P < 0.001) but only contributed 6.6% to the variance. Genetic distance (PhiST) calculated with AMOVA and multidimensional scaling (MDS) revealed that the South American and the Central American/Caribbean genotypes formed two separate clusters. East African varieties, which have unique characteristics from other traditional varieties, were distinct from other traditional varieties from South America and Oceania. These results support the reported hypothesis of the origin and dispersal of the sweetpotato and indicate that the primary centre of diversity probably has two distinct genepools. It is proposed that the dispersal of the sweetpotato from its origin may have mainly involved varieties from Central America/Caribbean as opposed to varieties from South America. There is an indication that new genepools may be evolving in Africa and Asia due to hybridisation and adaptation to the local environments.     

Identifying and selecting for genetic diversity in Papua New Guinea sweetpotato Ipomoea batatas (L.) Lam. germplasm collected as botanical seed

A total of 141 Ipomoea batatas (L.) Lam. accessionsderived from botanical seed originally collected from 26 sites in 4 Provinces inPapua New Guinea, a secondary center of genetic diversity for sweetpotato, weregenetically analyzed. Two hundred Amplified Fragment Length Polymorphism (AFLP)markers were identified and utilized in the analysis. Relatedness amongaccessions was estimated by analyzing the AFLP data using the Dice coefficientof similarity and UPGMA methods. The molecular analysis revealed relativelylimited genetic diversity within and between sites. Genotypes collected in agiven region often displayed molecular marker variability similar to thatobserved over the entire sampled area. However, a subset of 14 genotypes derivedfrom seed collected from New Ireland island differed from genotypes collected onNew Guinea island. Estimates of genetic diversity-based similarity valuescalculated from the AFLP data indicated a moderate level of diversity (0.767mean coefficient of similarity) across all plant materials analyzed. Threemethods of selection were evaluated for their efficacy in capturing themolecular marker diversity within the plant materials in the form of a subset.They were random, stratified-random (geographic based), and marker-assistedselection (MAS). MAS was the most efficient. A Maximally Diverse Subset (MDS) of12 genotypes capturing 92% of the molecular marker diversity was identified.     

Application of QUEFTS Model for Site-Specific Nutrient Management of NPK in Sweet Potato (Ipomoea batatas L. Lam)

Sweet potato productivity in India is either stagnated or lowering down over the past many years. The main reasons for low yield are conventional blanket recommendation of fertilizers, lower nutrient-use efficiency and imbalance in the use of fertilizers. Recommendation of major nutrients, nitrogen, phosphorus, and potassium (NPK), based on quantitative approaches will augment sweet potato production per unit area by increasing the nutrient-use efficiency. The present study calibrated the Quantitative Evaluation of Fertility of Tropical Soils (QUEFTS) model for the estimation of NPK requirements and fertilizers recommendations for different target yields of sweet potato. The QUEFTS basically works on the principle of NPK nutrient interactions and climate-adjusted yield potential of a region. Published data sets from several field experiments related to NPK carried out till date were collected to reflect the environment variability. The results of the present study showed that to produce one ton tuber, 16:6:18 kg N, P, and K, respectively, would be needed with the internal efficiencies of 61:167:57 kg tubers kg^?1 NPK removed. The maximum accumulation and dilution (kg tuber kg^?1 nutrient removed) of N (40, 80), P (96, 272), and K (30, 85) were also derived as standard parameters in QUEFTS for optimum fertilizer recommendation in tropical and subtropical regions of India. The observed yields of sweet potato with different amounts of nutrients were in agreement with the values predicted by the model. Therefore, it is utmost important to have the results of the study validated in major growing environments of India for fertilizer recommendation in sweet potato.     

Probing anthocyanin profiles in purple sweet potato cell line (Ipomoea batatas L. Cv. Ayamurasaki) by high-performance liquid chromatography and electrospray ionization tandem mass spectrometry

A purple line cell line (PL) generated from the storage root of purple-fleshed sweet potato (Ipomoea batatas L.) cv. Ayamurasaki produces a complex mixture of anthocyanins, and seven major anthocyanins have been isolated and identified to date. All these anthocyanins are exclusively cyanidin or peonidin 3-sophoroside-5-glucosides and their acylated derivatives. High-performance liquid chromatography (HPLC) coupled to photodiode array (PDA) detection and electrospray ionization tandem mass spectrometry (ESI-MS/MS) on a triple quadrupole instrument was employed to further investigate the anthocyanin composition of the PL extract. Precursor-ion analysis, product-ion analysis, and selected reaction monitoring (SRM) MS/MS experiments were conducted sequentially to screen and characterize anthocyanins in the aqueous extract of the PL cell line. Precursor-ion analysis specifically detected the molecular cations of each category of anthocyanins by scanning the precursors of anthocyanidins (cyanidin, peonidin, and pelargonidin). The detected molecular cation of each anthocyanin was fragmented using product-ion analysis by collisionally activated dissociation (CAD). MS/MS using SRM detection was conducted to further confirm the fragmentation observed during product-ion analysis. In comparison to the commonly used product-ion analysis technique, the combined use of precursor-ion analysis, product-ion analysis, and SRM is particularly useful for positive identification of anthocyanins in complex matrixes and provides important information to confirm the proposed structures. Twenty-six anthocyanins were detected and characterized in the aqueous extract of the PL cell line. Several anthocyanins, including two pelargonidin derivatives, were tentatively identified for the first time in these cells.