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Relationships in Ananas and other related genera using chloroplast DNA restriction site variation.

Chloroplast DNA (cpDNA) diversity was examined using PCR-RFLP to study phylogenetic relationships in Ananas and related genera. One hundred fifteen accessions representing the seven Ananas species and seven other Bromelioideae including the neighboring monospecific genus Pseudananas, two Pitcairnioideae, and one Tillandsioideae were included in the study. Eight primers designed from cpDNA were used for generating fragments. Restriction by 18 endonucleases generated 255 variable fragments. Dissimilarities were calculated from the resulting matrix using the Sokal and Michener index and the neighbor-joining method was used to reconstruct the diversity tree. Phylogenetic reconstruction was attempted using Wagner parsimony. Phenetic and cladistic analyses gave consistent results. They confirm the basal position of Bromelia in the Bromelioideae. Ananas and Pseudananas form a monophyletic group, with three strongly supported sub-groups, two of which are geographically consistent. The majority of Ananas parguazensis accessions constitute a northern group restricted to the Rio Negro and Orinoco basins in Brazil. The tetraploid Pseudananas sagenarius joins the diploid Ananas fritzmuelleri to constitute a southern group. The third and largest group, which includes all remaining species plus some accessions of A. parguazensis and intermediate phenotypes, is the most widespread and its distribution overlaps those of the northern and southern groups. Ananas ananassoides is dominant in this sub-group and highly variable. Its close relationship to all cultivated species supports the hypothesis that this species is the wild ancestor of the domesticated pineapple. The data indicate that gene flow is common within this group and scarcer with both the first and second groups. Comparison of cpDNA data with published genomic DNA data point to the hybrid origin of Ananas bracteatus and support the autopolyploidy of Pseudananas. The Ananas-Pseudananas group structure and distribution are consistent and we propose a scenario based on the refugia hypothesis to explain our data. These results and hypotheses bring some interesting points to consider in the current discussion on Ananas taxonomy.

Bromeliaceae↗

Gut colonization by an ice nucleation active bacterium, Erwinia (Pantoea) ananas reduces the cold hardiness of mulberry pyralid larvae.

To evaluate the suitability of using ice nucleation active (INA) bacteria for the biological control of insect pests, the supercooling point (SCP) of larvae of mulberry pyralid, Glyphodes duplicalis, and silkworm, Bombyx mori, ingesting INA strains of Erwinia (Pantoea) ananas and Pseudomonas syringae was determined. Mean SCP of the guts of silkworm larvae ingesting INA strains of E. ananas ranged from -2.5 to -2.8 degrees C, being 5 degrees C higher than that in control treatments. Similarly, mean SCP of mulberry pyralid larvae ingesting INA strain of E. ananas, which can grow well in the gut, was -4.7 degrees C at 3 days after treatment, being 6.5 degrees C higher than that in control treatments. On the other hand, mean SCP of the larvae-ingesting INA strain of P. syringae, which cannot grow in the gut, was -9.0 degrees C at 3 days after treatment, rising by only 2.5 degrees C higher than that in the control treatments. In addition, more than 80% of the larvae of mulberry pyralid ingesting the INA strain of E. ananas froze and eventually died when exposed to -6 degrees C for 18 h, while only 36% of the larvae ingesting the INA strain of P. syringae, or approximately 20% of the control larvae, froze and died. Thus, the gut colonization by INA strains of E. ananas reduced remarkably the cold hardiness of the insects. These findings suggest that INA strains of E. ananas could be effective as a potential biological control agent of insect pests.

Animals↗

The effects of Ananas comosus L. leaves on diabetic-dyslipidemic rats induced by alloxan and a high-fat/high-cholesterol diet.

The aim of this study is to demonstrate the effects of Ananas comosus L. leaves on diabetic-dyslipidemic rats. Hypoglycemic and hypolipidemic activities of the ethanolic extract of Ananas comosus L. leaves (EEACL) were evaluated in normal and alloxan-induced diabetic rats by oral glucose tolerance test and an olive oil load test. Anti-diabetic, anti-hyperlipidemic and anti-oxidative activities of EEACL were also investigated in diabetic-dyslipidemic rats induced by alloxan and a high-fat/high-cholesterol diet. EEACL at the dose of 0.40 g/kg significantly inhibited the increase in blood glucose in diabetic rats in oral glucose tolerance test, but did not cause any hypoglycerimic activity in normal rats. It also significantly inhibited the increase in postprandial triglycerides (TG) levels in both normal and diabetic rats in olive oil load test. After 15 days of treatment of diabetic dyslipidemic rats, EEACL significantly decreased blood glucose (-51.0%, P < 0.01), TG (-50.1%, P < 0.01), TC (-23.3%, P < 0.01), LDL-c (-47.9%, P < 0.01) and glycated albumin (-25.4%, P < 0.01) levels, significantly increased serum high-density lipoprotein cholesterol levels (66.2%, P < 0.01) and prevented lower body weight of diabetes (11.8%, P < 0.05), significantly lowered lipid peroxidation productions of blood (-27.8%, P < 0.01), brain (-31.6%, P < 0.05), liver (-44.5%, P < 0.01) and kidneys (-72.2%, P < 0.05) compared with those in untreated diabetic dyslipidemic rats. These data suggest that EEACL has anti-diabetic, anti-dyslipidemic and anti-oxidative activities, which may be developed into a new plant medicine for treatment of diabetes and its complications.

Ananas↗

Retrotransposon-like sequences integrated into the genome of pineapple, Ananas comosus.

Retrotransposon-like sequences have been serendipitously detected in the genome of commercial pineapple, Ananas comosus. The sequence from a 2.6 kb cloned fragment of this element had greatest similarity to the del1 Lilium henryi retrotransposon and the gypsy/Ty3 group of retroelements. The order of the genes from 5' to 3' was reverse transcriptase, ribonuclease H and integrase. The integrase domain contained the amino acid sequence motifs which have been associated with recognition of the long terminal repeats and with the cutting/joining reactions required for integration of similar retroelements into the host genome. The retrotransposon existed as a population of variable sequences which were dispersed throughout the genome of pineapple. Southern hybridisation showed that the retrotransposon had integrated repeatedly into the pineapple genome. The reading frame of the element was not interrupted by stop codons, suggesting that it is still potentially capable of transposing. This is the first report of a retrotransposon in pineapple, which we have called deal (for dispersed element of Ananas).

Amino Acid Sequence↗

Pharmacokinetic study of p-coumaric acid in mouse after oral administration of extract of Ananas comosus L. leaves.

Quantification of p-coumaric acid in mouse plasma following oral administration of Ananas comosus L. leaves was achieved by reversed-phase high-performance liquid chromatography using a mobile phase of water-acetonitrile (82:18, v/v) and UV detection at 310 nm. The method was linear (determination coefficient, r2 = 0.9997) within the tested range (0.04-1.28 microg/mL). Intra- and inter-day precision coefficients of variation and accuracy bias were acceptable (maximal CV value was 4.06% for intra-day and 4.19% for inter-day) over the entire range. The recoveries were 90.63, 97.98 and 100.01% for concentrations of 0.04, 0.32 and 1.28 microg/mL, respectively. This is a very rapid, sensitive and economical way to determine p-coumaric acid concentration in mouse plasma after oral administration of A. comosus leaves. The concentration-time curve was fitted to the one-compartment model. This is the first time that p-coumaric acid extracted from A. comosus leaves was detected by HPLC-UV method and its pharmacokinetic characteristic was comprehensively studied.

Administration, Oral↗

Effect of ethanolic extracts of Ananas comosus L. leaves on insulin sensitivity in rats and HepG2.

Ethanolic extracts of Ananas comosus L. leaves (AC) enriched with phenols have hypoglycemic activity in diabetic rats. Here, we investigated the effect of AC on insulin sensitivity in rats and HepG2. In high-fat diet-fed and low-dose streptozotozin-treated diabetic Wistar rats subjected to challenge with exogenous human insulin, AC treatment at an oral dose of 0.40 g/kg could significantly improve sensitivity to exogenous insulin. After a sub-acute treatment, AC also could inhibit the development of insulin resistance in high-fat diet-fed and low-dose streptozotozin-treated diabetic rats following the test of loss of tolbutamide-induced blood glucose lowering action. For intravenous insulin/glucose infusion test, high-fat diet-fed and low-dose alloxan-treated Wistar rats were associated with insulin resistance, which was improved after AC or fenofibrate treatment. AC application inhibited the development of insulin resistance in HepG2 cells. The above animal models were well developed to simulate type 2 diabetes. Taken together, our results suggest that AC may improve insulin sensitivity in type 2 diabetes and could be developed into a new potential natural product for handling of insulin resistance in diabetic patients.

Ananas↗

On-line gas chromatography combustion/pyrolysis isotope ratio mass spectrometry (HRGC-C/P-IRMS) of pineapple (Ananas comosus L. Merr.) volatiles.

By use of extracts prepared by liquid-liquid separation of the volatiles from self-prepared juices of pineapple fruits (Ananas comosus) (n = 14) as well as commercial pineapple recovery aromas/water phases (n = 3), on-line capillary gas chromatography-isotope ratio mass spectrometry was employed in the combustion (C) and the pyrolysis (P) modes (HRGC-C/P-IRMS) to determine the delta(13)C(VPDB) and delta(2)H(VSMOW) values of selected pineapple flavor constituents. In addition to methyl 2-methylbutanoate 1, ethyl 2-methylbutanoate 2, methyl hexanoate 3, ethyl hexanoate 4, and 2,5-dimethyl-4-methoxy-3[2H]-furanone 5, each originating from the fruit, the delta(13)C(VPDB) and delta(2)H(VSMOW) data of commercial synthetic 1-5 and "natural" (biotechnologically derived) 1-4 were determined. With delta(13)C(VPDB) data of pineapple volatiles 1-4 varying from -12.8 to -24.4 per thousand, the range expected for CAM metabolism was observed. Compound 5 showed higher depletion from -20.9 to -28.6 per thousand. A similar situation was given for the delta(2)H(VSMOW) values of 3-5 from pineapple ranging from -118 to -191 per thousand, whereas 1 and 2 showed higher depleted values from -184 to -263 per thousand. In nearly all cases, analytical differentiation of 1-5 from pineapple and natural as well as synthetic origin was possible. In general, natural and synthetic 1-5 exhibited delta(13)C(VPDB) data ranging from -11.8 to -32.2 per thousand and -22.7 to -35.9 per thousand, respectively. Their delta(2)H(VSMOW) data were in the range from -242 to -323 per thousand and -49 to -163 per thousand, respectively.

Ananas↗

Respiratory properties and malate metabolism in Percoll-purified mitochondria isolated from pineapple, Ananas comosus (L.) Merr. cv. smooth cayenne.

An investigation was made of the respiratory properties and the role of the mitochondria isolated from one phosphoenolpyruvate carboxykinase (PCK)-CAM plant Ananas comosus (pineapple) in malate metabolism during CAM phase III. Pineapple mitochondria showed very high malate dehydrogenase (MDH), and low malic enzyme (ME) and glutamate-oxaloacetate transaminase (GOT) activities. The mitochondria readily oxidized succinate and NADH with high rates and coupling, while they only oxidized NADPH in the presence of Ca(2+). Pineapple mitochondria oxidized malate with low rates under most assay conditions, despite increasing malate concentrations, optimizing pH, providing cofactors such as coenzyme A, thiamine pyrophosphate, and NAD(+), and supplying individually external glutamate or GOT. However, providing glutamate and GOT simultaneously strongly increased the rates of malate oxidation. The OAA easily permeated the mitochondrial membranes to import into or export out of pineapple mitochondria during malate oxidation, but the mitochondria did not consume external Asp or alpha-KG. These results suggest that OAA played a significant role in the mitochondrial malate metabolism of pineapple, in which malate was mainly oxidized by active mMDH to produce OAA which could be exported outside the mitochondria via a malate-OAA shuttle. Cytosolic GOT then consumed OAA by transamination in the presence of glutamate, leading to a large increase in respiration rates. The malate-OAA shuttle might operate as a supporting system for decarboxylation in phase III of PCK-CAM pineapple. This shuttle system may be important in pineapple to provide a source of energy and substrate OAA for cytosolic PCK activity during the day when cytosolic OAA and ATP was limited for the overall decarboxylation process.

Ananas↗

Proliferation potential of 18-month-old callus of Ananas comosus L. cv. Moris.

Differential effect of plant growth regulators and additives in proliferation of 18-month-old calli of Ananas comosus L. cv. Moris were assessed in vitro. The proliferation of callus relied on the growth regulators and additives. Of the different auxins supplemented in the Murashige and Skoog (MS) media, 32.22 microM alpha-naphthaleneacetic acid (NAA) gave the highest mean fresh weight of callus (46.817 g). Medium supplemented with 2,4-dichlorophenoxyacetic acid (2,4-D) was inferior to NAA, while b-naphthoxy acetic acid (BNOA) and p-chlorophenoxy acetic acid (4-CPA) were not effective in proliferating 18-months old callus. Addition of casein hydrolysate and coconut water to NAA supplemented medium showed better proliferation and production of callus. However, in terms of callus production, NAA at 32.22 microM was economically better.

Ananas↗

Purification, characterization, and antifungal activity of chitinases from pineapple (Ananas comosus) leaf.

Three chitinases, designated pineapple leaf chitinase (PL Chi)-A, -B, and -C were purified from the leaves of pineapple (Ananas comosus) using chitin affinity column chromatography followed by several column chromatographies. PL Chi-A is a class III chitinase having a molecular mass of 25 kDa and an isoelectric point of 4.4. PL Chi-B and -C are class I chitinases having molecular masses of 33 kDa and 39 kDa and isoelectric points of 7.9 and 4.6 respectively. PL Chi-C is a glycoprotein and the others are simple proteins. The optimum pHs of PL Chi-A, -B, and -C toward glycolchitin are pH 3, 4, and 9 respectively. The chitin-binding ability of PL Chi-C is higher than that of PL Chi-B, and PL Chi-A has lower chitin-binding ability than the others. At low ionic strength, PL Chi-B exhibits strong antifungal activity toward Trichoderma viride but the others do not. At high ionic strength, PL Chi-B and -C exhibit strong and weak antifungal activity respectively. PL Chi-A does not have antifungal activity.

Amino Acid Sequence↗

Pineapple [Ananas comosus (L.) Merr].

A procedure for pineapple [Ananas comosus (L.) Merr.] genetic transformation is described, which involves temporary immersion bioreactors (TIB) for selection of transgenic plants. Success in the production of transgenic pineapple plants combines tissue culture factors. Firstly, the use of regenerable pineapple callus as starting material for transformation whose cells shown to be competent for Agrobacterium infection. Secondly, the used of filtered callus, resulting in homogeneously sized clusters, thereby increasing the contact between the cell surfaces and A. tumefaciens and releasing phenolic compounds which induce Agrobacterium virulence. Thirdly, regeneration of primary plants without selection pressure, that allowing a massive production of putative transgenic pineapples. Finally, we support that TIB technology is a powerful system to recover nonchimera transgenic plants by micropropagation with the use of an adequate selection agent.

Agrobacterium tumefaciens↗

Routes of Ethephon Uptake in Pineapple (Ananas comosus) and Reasons for Failure of Flower Induction.

Ethylene-releasing agents such as ethephon (2-chloroethylphosphonic acid) are used widely to induce flowering in pineapples (Ananas comosus (L.) Merrill). However, ethephon treatment is less reliable in summer, particularly if plants are treated on abnormally hot days. [(14)C]ethephon was used to follow uptake and translocation in leaf tissues. Up to 30% of the ethephon entered the leaf within 4 h, and up to 60% by 24 h. Uptake was dramatically modified by temperature, relative humidity, solution pH, and the surface on which solution droplets were placed. Entry occurred across the leaf cuticle and probably also by way of stomatal pores, and label was recovered at all depths within the leaf. (14)C label entered more rapidly through the abaxial epidermis than through the adaxial epidermis. Low-volume spray applications to whole plants resulted in rapidly drying droplets mainly on the adaxial, distal epidermis and were rather ineffective at inducing flowering, possibly because little ethephon or ethylene reaches the shoot apex. High-volume sprays may facilitate ethephon entry because solution accumulates in leaf axils and hence remains in prolonged contact with abaxial epidermis of leaf bases close to the shoot apex. When poured into the center of the plant, 20% of a normal commercial ethephon dose induced full flowering even under adverse temperatures. It is suggested that high-volume evening spraying and avoidance of hot days may reduce the incidence of flowering failure.

Journal Article↗

An ice nucleation active gene of Erwinia ananas. Sequence similarity to those of Pseudomonas species and regions required for ice nucleation activity.

The ice nucleation active gene, inaA, of Erwinia ananas IN-10 has been sequenced. This gene encodes a protein composed of 1322 amino acid residues. The inaA protein contains a 1120-residue segment consisting of 70 repeats of closely related 16 amino acid motifs (R-domain), which is flanked by N- and C-terminal sequences (N- and C-domains, respectively). Its primary structure is similar to, but not identical with, those of Pseudomonas inaW and inaZ gene products. By truncating the inaA gene to various extents, it was found that deletion of the C-domain resulted in complete loss of the ice nucleation activity, whereas removal of the N-domain led to a moderate decrease in the activity. Complete loss of the activity was also observed when the N-domain plus a large part of the P-domain were deleted. It is suggested that the C-domain is required for the assembly of inaA protein to form a functional ice nucleus.

Amino Acid Sequence↗

Characterization of sinapyl derivatives in pineapple (Ananas comosus [L.] Merill) juice.

Three previously unidentified phenolic compounds were found in pineapple (Ananas comosus [L.] Merill) juice in substantial concentrations and were isolated by semipreparative reverse phase HPLC. The structures were elucidated from UV spectra, acid hydrolysis, and subsequent amino acid analysis, mass spectrometry, and two-dimensional NMR spectroscopy. The compounds are identified as S-sinapyl-L-cysteine, N-L-gamma-glutamyl-S-sinapyl-L-cysteine, and S-sinapylglutathione.

Beverages↗

A comparative study on diurnal changes in metabolite levels in the leaves of three crassulacean acid metabolism (CAM) species, Ananas comosus, Kalanchoë daigremontiana and K. pinnata.

A comparative study on diurnal changes in metabolite levels associated with crassulacean acid metabolism (CAM) in the leaves of three CAM species, Ananas comosus (pineapple), a hexose-utilizing species, and Kalanchoë daigremontiana and K. pinnata, two starch-utilizing species, were made. All three CAM species showed a typical feature of CAM with nocturnal malate increase. In the two Kalanchoë species, isocitrate levels were higher than citrate levels; the reverse was the case in pineapple. In the two Kalanchoë species, a small nocturnal citrate increase was found and K. daigremontiana showed a small nocturnal isocitrate increase. Glucose 6-phosphate (G-6-P), fructose 6-phosphate (F-6-P) and glucose 1-phosphate (G-1-P) levels in the three CAM species rose rapidly during the first part of the dark period and decreased during the latter part of the dark period. The levels of the metabolites also decreased during the first 3 h of the light period, then, remained little changed through the rest of the light period. Absolute levels of G-6-P, F-6-P and G-1-P were higher in pineapple than in the two Kalanchoë species. Fructose 1,6-bisphosphate (F-1,6-P(2)) levels in the three CAM species increased during the dark period, then dramatically decreased during the first 3 h of the light period and remained unchanged through the rest of the light period. The extent of nocturnal F-1,6-P(2) increase was far greater in the two Kalanchoë species than in pineapple. Absolute levels of F-1,6-P(2) were higher in the two Kalanchoë species than in pineapple, especially during dark period. Diurnal changes in oxaloacetate (OAA), pyruvate (Pyr) and phosphoenolpyruvate (PEP) levels in the three CAM species were similar.

Carboxylic Acids↗

Polysaccharide composition of unlignified cell walls of pineapple [Ananas comosus (L.) Merr.] fruit.

The polysaccharides of cell walls isolated from the fleshy, edible part of the fruit of the monocotyledon pineapple [Ananas comosus (L.) Merr.] (family Bromeliaceae) were analyzed chemically. These cell walls were derived mostly from parenchyma cells and were shown histochemically to be unlignified, but they contained ester-linked ferulic acid. The analyses indicated that the noncellulosic polysaccharide composition of the cell walls was intermediate between that of unlignified cell walls of species of the monocotyledon family Poaceae (grasses and cereals) and that of unlignified cell walls of dicotyledons. Glucuronoarabinoxylans were the major non-cellulosic polysaccharides in the pineapple cell walls. Xyloglucans were also present, together with small amounts of pectic polysaccharides and glucomannans (or galactoglucomannans). The large amounts of glucuronoarabinoxylans and small amounts of pectic polysaccharides resemble the noncellulosic polysaccharide composition of the unlignified cell walls of the Poaceae. However, the absence of (1-->3,1-->4)-beta-glucans, the presence of relatively large amounts of xyloglucans, and the possible structure of the xyloglucans resemble the noncellulosic polysaccharide composition of the unlignified cell walls of dicotyledons.

Carbohydrate Sequence↗

Carbon Dioxide Exchange and Acidity Levels in Detached Pineapple, Ananas comosus (L.), Merr., Leaves during the Day at Various Temperatures, Oxygen and Carbon Dioxide Concentrations.

The effects of temperature, O(2), and CO(2) on titratable acid content and on CO(2) exchange were measured in detached pineapple (Ananas comosus) leaves during the daily 15-hour light period. Comparative measurements were made in air and in CO(2)-free air. Increasing the leaf temperature from 20 to 35 C decreased the total CO(2) uptake in air and slightly increased the total CO(2) released into CO(2)-free air. Between 25 and 35 C, the activation energy for daily acid loss was near 12 kcal mol(-1), but at lower temperatures the activation energy was much greater.Increasing O(2) or decreasing the CO(2) concentration decreased the total CO(2) fixation in air, whereas the total CO(2) released in CO(2)-free air was increased. The total acid content remained constant at 20 C, but it decreased progressively with increasing temperature both in air and in CO(2)-free air. The total acid content at 30 C remained constant in 2% O(2) irrespective of CO(2) concentration. The total acid content decreased in 21 and 50% O(2) as the CO(2) increased from 0 to 300, and 540 mul/l of CO(2). The data indicate that photorespiration is present in pineapple. The lack of acid loss in 2% O(2) suggests that light deacidification is dependent upon respiration and that higher O(2) concentrations are required to saturate deacidification.

Journal Article↗

Characterization of Phosphoenolpyruvate Carboxykinase from Pineapple Leaves Ananas comosus (L.) Merr.

Phosphoenolpyruvate carboxykinase has been partially purified from pineapple (Ananas comosus [L.]) leaves. Specific activities obtained show it to be a major activity in this tissue. Above 15 C, the respective activation energies for decarboxylation and carboxylation are 13 and 12 kcal/mol. Below 15 C, there are discontinuities in Arrhenius plots with an associated large increase in activation energy. The adenine nucleotides are preferred to other nucleotides as substrates. The apparent Km values in the carboxylation direction are: ADP 0.13 mm, HCO(3) (-) 3.4 mm, and phosphoenolpyruvate 5 mm. In the decarboxylation direction, the apparent Km values are: ATP 0.02 mm, ADP 0.05 mm, and oxaloacetate 0.4 mm. The decarboxylation activity had an almost equal velocity with either ADP or ATP. The pH optima are between 6.8 and 7. Inhibition of the carboxylation reaction by ATP, pyruvate, and carbonic anhydrase was demonstrated. Decarboxylase specific activities are over twice carboxylation activities. The data support a model in which phosphoenolpyruvate carboxykinase is of physiological significance only during the light period and then only as a decarboxylase.

Journal Article↗