phyllocactin and hylocerenin

Betacyanins are reddish to violet pigments that can be found in red pitahaya (Hylocereus polyrhizus) and red spinach (Amaranthus dubius). This study investigated the impact of sub-fractionation (solvent partitioning) on betacyanin content in both plants. Characterization of betacyanins and evaluation of their antimicrobial activities were also carried out. Betanin was found in both plants. In addition, isobetanin, phyllocactin and hylocerenin were found in red pitahaya whereas amaranthine and decarboxy-amaranthine were found in red spinach. Sub-fractionated red pitahaya and red spinach had 23.5 and 121.5 % more betacyanin content, respectively, than those without sub-fractionation. Sub-fractionation increased the betanin and decarboxy-amaranthine content in red pitahaya and red spinach, respectively. The betacyanin fraction from red spinach (minimum inhibitory concentration [MIC] values: 0.78-3.13 mg/mL) demonstrated a better antimicrobial activity profile than that of red pitahaya (MIC values: 3.13-6.25 mg/mL) against nine Gram-positive bacterial strains. Similarly, the red spinach fraction (MIC values: 1.56-3.13 mg/mL) was more active than the red pitahaya fraction (MIC values: 3.13-6.25 mg/mL) against five Gram-negative bacterial strains. This could be because of a higher amount of betacyanin, particularly amaranthine in the red spinach.

Topics: Amaranthus; Anti-Infective Agents; Betacyanins; Cactaceae; Chromatography, High Pressure Liquid; Plant Extracts

Polar betacyanin pigments together with betaxanthins from ripe cactus fruits of Hylocereus polyrhizus (Cactaceae) were fractionated by means of preparative ion-pair high-speed countercurrent chromatography (IP-HSCCC) also using the elution-extrusion (EE) approach for a complete pigment recovery. HSCCC separations were operated in the classical 'head-to-tail' mode with an aqueous mobile phase. Different CCC solvent systems were evaluated in respect of influence and effectiveness of fractionation capabilities to separate the occurring pigment profile of H. polyrhizus. For that reason, the additions of two different volatile ion-pair forming perfluorinated carboxylic acids (PFCA) were investigated. For a direct comparison, five samples of Hylocereus pigment extract were run on preparative scale (900 mg) in 1-butanol-acetonitrile-aqueous TFA 0.7% (5:1:6, v/v/v) and the modified systems tert.-butyl methyl ether-1-butanol-acetonitrile-aqueous PFCA (2:2:1:5, v/v/v/v) using 0.7% and 1.0% trifluoroacetic acid (TFA) or heptafluorobutyric acid (HFBA) in the aqueous phase, respectively. The chemical affinity to the organic stationary CCC solvent phases and in consequence the retention of these highly polar betalain pigments was significantly increased by the use of the more lipophilic fluorinated ion-pair reagent HFBA instead of TFA. The HFBA additions separated more effectively the typical cacti pigments phyllocactin and hylocerenin from betanin as well as their iso-forms. Unfortunately, similar K(D) ratios and selectivity factors alpha around 1.0-1.1 in all tested solvent systems proved that the corresponding diastereomers, 15S-type pigments cannot be resolved from the 15R-epimers (iso-forms). Surprisingly, additions of the stronger ion-pair reagent (HFBA) resulted in a partial separation of hylocerenin from phyllocactin which were not resolved in the other solvent systems. The pigments were detected by means of HPLC-DAD and HPLC-electrospray ionization-MS using also authentic reference materials.

Topics: Analytic Sample Preparation Methods; Artifacts; Betacyanins; Betalains; Betaxanthins; Cactaceae; Chromatography, High Pressure Liquid; Countercurrent Distribution; Fluorocarbons; Fruit; Methyl Ethers; Plant Extracts; Solvents; Spectrometry, Mass, Electrospray Ionization; Trifluoroacetic Acid

A method was developed for identification of diastereomers of 2-decarboxy-betacyanins and 2,17-bidecarboxy-betacyanins chromatographed in reversed-phase high-performance liquid chromatography (HPLC) as pairs of unknown elution order. The method was based on alkaline hydrolysis of selected betacyanin and decarboxylated betacyanin mixtures and subsequent cross-recondensation of the hydrolysates. The arising intermediate derivatives of decarboxylated betalamic acid and cyclo-dopa were stable enough for subsequent recondensation. Generated diagnostic pigments as the recondensation products were monitored by HPLC-diode-array detection-electrospray ionisation mass spectrometry. The isoforms (15R) of 2-decarboxy-betacyanins and 2,17-bidecarboxy-betacyanins were eluted earlier than the 15S forms, in contrast to betacyanins and 17-decarboxy-betacyanins, but in accordance with betaxanthins. 2,17-Bidecarboxy-betanin/2,17-bidecarboxy-isobetanin, being not resolved in reversed-phase HPLC, were partially separated by ion-pair chromatography and under these conditions their order of elution was the same as that of acylated 2,17-bidecarboxy-betacyanins. The method allows complete identification of all the decarboxylated groups of betanin, phyllocactin and hylocerenin as well as other betacyanins in biological or pharmaceutical material.

Topics: Betacyanins; Chromatography, High Pressure Liquid; Methods; Spectrometry, Mass, Electrospray Ionization; Stereoisomerism

Thermal degradation of betanin, phyllocactin (malonyl-betanin), and hylocerenin (3' '-hydroxy-3' '-methyl-glutaryl-betanin) solutions isolated from purple pitaya (Hylocereus polyrhizus [Weber] Britton and Rose) was monitored by spectrophotometric and high-performance liquid chromatography-diode array detection (HPLC-DAD) analyses. For betanin and phyllocactin solutions, the color shift upon thermal treatment was found to be nearly identical, while hylocerenin samples exhibited an intelligibly higher chromatic steadiness. Betanin proved to be the most stable individual pigment structure, while the enhanced tinctorial stability of the integral phyllocactin and especially hylocerenin solutions was due to the formation of red degradation products exhibiting improved color retention as opposed to their respective genuine pigments. Individual structure-related stability characteristics can exclusively be assessed by HPLC-DAD analyses and may not be noticed by mere spectrophotometric assessment of color and tinctorial strength.

Topics: Betacyanins; Cactaceae; Chromatography, High Pressure Liquid; Color; Coloring Agents; Drug Stability; Hot Temperature; Solutions; Spectrophotometry

Betanin, phyllocactin (malonylbetanin) and hylocerenin (3-hydroxy-3-methylglutarylbetanin) were isolated from purple pitaya (Hylocereus polyrhizus [Weber] Britton and Rose) juice, and their degradation products generated by heating at 85 degrees C were subsequently monitored by high-performance liquid chromatography/electrospray ionization tandem mass spectrometry. Thermal degradation of phyllocactin and hylocerenin in purified solution excluding the alleged protective effects by the juice matrix is reported for the first time. Betanin was predominantly degraded by hydrolytic cleavage, while decarboxylation and dehydrogenation were of minor relevance. In contrast, hylocerenin showed a strong tendency to decarboxylation and dehydrogenation, hydrolytic cleavage of the aldimine bond occurring secondarily. Phyllocactin degradation was most complex because of additional decarboxylation of the malonic acid moiety as well as generation and subsequent degradation of betanin due to phyllocactin demalonylation. Upon prolonged heating, all betacyanins under observation formed degradation products characterized by an additional double bond at C2-C3. Hydrolytic cleavage of the aldimine bond of phyllocactin and hylocerenin yielded previously unknown acylated cyclo-dopa derivatives traceable by positive ionization, while application of ESI(-) facilitated the detection of a glycosylated aminopropanal derivative and dopamine, which have never been described before as betanin degradation products.

Topics: Betacyanins; Chromatography, High Pressure Liquid; Hot Temperature; Molecular Structure; Plant Extracts; Spectrometry, Mass, Electrospray Ionization