Carbon flow and carbohydrate metabolism during sink-to-source transition for developing cladodes of Opuntia ficus-indica

Daughter cladodes (flattened stem segments) of Opuntia ficus-indica (L.) Miller at 14-18 d after appearance on the underlying basal cladodes were sinks, requiring carbohydrate import for growth. Import stopped at 25-36 d, and the daughter cladodes became sources at 27-28 d. The activities of Rubisco, PEPCase, and sucrose-Pi synthase as well as the chlorophyll content at 14 d were not less than those at 28 d, suggesting that photosynthetic or sucrose synthesis capacity was not limiting carbon assimilation for sink cladodes. Sucrose synthase (SS) activity was three times higher than that of alkaline invertase, indicating that SS is the major enzyme for cytoplasmic sucrose hydrolysis. The SS activity was correlated with cladode growth, the highest activity coinciding with the highest growth rate. The sink-to-source transition for daughter cladodes was correlated with increases in malate and H⁺ concentrations in the vacuoles of chlorenchyma cells, with 5-fold higher nocturnal malate production and 10-fold higher H⁺ concentration in 28- than in 14-d-old daughter cladodes. The vacuolar H⁺ increase during cladode development would lower cytoplasmic pH, which may trigger metabolic events affecting the sink-to-source transition.     

Effects of drought on water relations and nonstructural carbohydrates in cladodes of Opuntia ficus-indica

Nocturnal acid accumulation, water content, osmotic pressure (π), and nonstructural carbohydrates were determined in the chlorenchyma and the water-storage parenchyma of Opuntia ficus-indica (L.) Miller for well-watered plants and those subjected to drought for 15 weeks. During the 15-week drought, total cladode water content decreased by 57%, the water-storage parenchyma losing a greater fraction of water than the chlorenchyma, which most likely helped maintain nocturnal acid accumulation in the latter tissue. Despite the preferential water loss from the water-storage parenchyma, it had a lower π than the chlorenchyma over the 15 weeks of drought, suggesting a substantial decrease in osmotically active solutes in the water-storage parenchyma. Also, the measured π increases of both tissues were much less than those predicted based on the loss of water during drought and the initial content of osmotically active solutes under well-watered conditions. A decrease in the amount of soluble sugars (glucose. fructose and sucrose) occurred in plants subjected to drought. accounting for 46% and 81% of the difference between the measured and the predicted increases in π of the chlorenchyma and the water-storage parenchyma. respectively. The decrease in soluble sugars was associated with an equivalenl increase in polysaccharides, presumably starch, in the water-storage parenchyma. but not in the chlorenchyma.     

Young daughter cladodes affect CO2 uptake by mother cladodes of Opuntia ficus-indica

BACKGROUND AND AIMS: Drought damages cultivated C3, C4 and CAM plants in the semi-arid lands of central Mexico. Drought damage to Opuntia is common when mother cladodes, planted during the dry spring season, develop young daughter cladodes that behave like C3 plants, with daytime stomatal opening and water loss. In contrast, wild Opuntia are less affected because daughter cladodes do not develop on them under extreme drought conditions. The main objective of this work is to evaluate the effects of the number of daughter cladodes on gas exchange parameters of mother cladodes of Opuntia ficus-indica exposed to varying soil water contents. METHODS: Rates of net CO2 uptake, stomatal conductance, intercellular CO2 concentration, chlorophyll content and relative water content were measured in mature mother cladodes with a variable number of daughter cladodes growing in spring under dry and wet conditions. KEY RESULTS: Daily carbon gain by mother cladodes was reduced as the number of daughter cladodes increased to eight, especially during drought. This was accompanied by decreased mother cladode relative water content, suggesting movement of water from mother to daughter cladodes. CO2 assimilation was most affected in phase IV of CAM (late afternoon net CO2 uptake) by the combined effects of daughter cladodes and drought. Rainfall raised the soil water content, decreasing the effects of daughter cladodes on net CO2 uptake by mother cladodes. CONCLUSIONS: Daughter cladodes significantly hasten the effects of drought on mother cladodes by competition for the water supply and thus decrease daily carbon gain by mother cladodes, mainly by inhibiting phase IV of CAM.     

Protective effect of Opuntia ficus-indica L. cladodes against UVA-induced oxidative stress in normal human keratinocytes

Opuntia ficus-indica L. is known for its beneficial effects on human health, but still little is known on cladodes as a potent source of antioxidants. Here, a direct, economic and safe method was set up to obtain water extracts from Opuntia ficus-indica cladodes rich in antioxidant compounds. When human keratinocytes were pre-treated with the extract before being exposed to UVA radiations, a clear protective effect against UVA-induced stress was evidenced, as indicated by the inhibition of stress-induced processes, such as free radicals production, lipid peroxidation and GSH depletion. Moreover, a clear protective effect against apoptosis in pre-treated irradiated cells was evidenced. We found that eucomic and piscidic acids were responsible for the anti-oxidative stress action of cladode extract. In conclusion, a bioactive, safe, low-cost and high value-added extract from Opuntia cladodes was obtained to be used for skin health/protection.     

Opuntia ficus-indica cladodes as feedstock for ethanol production by Kluyveromyces marxianus and Saccharomyces cerevisiae

The feasibility of ethanol production using an enzymatic hydrolysate of pretreated cladodes of Opuntia ficus-indica (prickly pear cactus) as carbohydrate feedstock was investigated, including a comprehensive chemical analysis of the cladode biomass and the effects of limited aeration on the fermentation profiles and sugar utilization. The low xylose and negligible mannose content of the cladode biomass used in this study suggested that the hemicellulose structure of the O. ficus-indica cladode was atypical of hardwood or softwood hemicelluloses. Separate hydrolysis and fermentation and simultaneous saccharification and fermentation procedures using Kluyveromyces marxianus and Saccharomyces cerevisiae at 40 and 35 °C, respectively, gave similar ethanol yields under non-aerated conditions. In oxygen-limited cultures K. marxianus exhibited almost double the ethanol productivity compared to non-aerated cultures, although after sugar depletion utilization of the produced ethanol was evident. Ethanol concentrations of up to 19.5 and 20.6 g l^?1 were obtained with K. marxianus and S. cerevisiae, respectively, representing 66 and 70 % of the theoretical yield on total sugars in the hydrolysate. Because of the low xylan content of the cladode biomass, a yeast capable of xylose fermentation might not be a prerequisite for ethanol production. K. marxianus, therefore, has potential as an alternative to S. cerevisiae for bioethanol production. However, the relatively low concentration of fermentable sugars in the O. ficus-indica cladode hydrolysate presents a technical constraint for commercial exploitation.     

Effects of shade, drought and daughter cladodes on the CO2 uptake by cladodes of Opuntia ficus-indica

The effects of shade on the physiology of opuntias have received little attention, notwithstanding that shade regularly occurs in both wild stands and cultivated populations. This research evaluates the effects of shade on the physiology of cladodes of Opuntia ficus‐indica, with and without daughter cladodes, as they are exposed to progressive drought. The stress caused by shade, drought and daughter cladodes reduced photosynthesis by mother cladodes and was associated with decreases in relative water content, parenchyma thickness and chlorophyll content. Shade exacerbated the physiological drought of mother cladodes imposed by daughter cladodes and by reduced soil water content.     

The mucilage of Opuntia ficus-indica

The mucilage extracted from the cladodes (modified stems) of Opuntia ficus-indica contains residues of d-galactose, d-xylose, l-arabinose, l-rhamnose, and d-galacturonic acid. Seasonal variation in the sugar composition of the mucilage has been investigated. Fractionation studies indicate that the mucilage is essentially homogeneous. The rate of release of the constituent sugars and the change in optical rotation on mild hydrolysis coupled with the results of chromic acid oxidation suggest that the mucilage contains α-arabinofuranosyl, β-xylopyranosyl, β-rhamnopyranosyl, β-galactopyranosyl, and α-galactopyranosyluronic acid residues. The results also suggest a core containing galacturonic acid, rhamnose, and galactose, to which xylose and arabinose are attached in peripheral positions.     

Metabolic profiling of the sink-to-source transition in developing leaves of quaking aspen

Profiles of small polar metabolites from aspen (Populus tremuloides Michx.) leaves spanning the sink-to-source transition zone were compared. Approximately 25% of 250 to 300 routinely resolved peaks were identified, with carbohydrates, organic acids, and amino acids being most abundant. Two-thirds of identified metabolites exhibited greater than 4-fold changes in abundance during leaf ontogeny. In the context of photosynthetic and respiratory measurements, profile data yielded information consistent with expected developmental trends in carbon-heterotrophic and carbon-autotrophic metabolism. Suc concentration increased throughout leaf expansion, while hexose sugar concentrations peaked at mid-expansion and decreased sharply thereafter. Amino acid contents generally decreased during leaf expansion, but an early increase in Phe and a later one in Gly and Ser reflected growing commitments to secondary metabolism and photorespiration, respectively. The assimilation of nitrate and utilization of stored Asn appeared to be marked by sequential changes in malate concentration and Asn transaminase activity. Principal component and hierarchical clustering analysis facilitated the grouping of cell wall maturation (pectins, hemicelluloses, and oxalate) and membrane biogenesis markers in relation to developmental changes in carbon and nitrogen assimilation. Metabolite profiling will facilitate investigation of nitrogen use and cellular development in Populus sp. varying widely in their growth and pattern of carbon allocation during sink-to-source development and in response to stress.     

Characterization of crystalline structures in Opuntia ficus-indica

This research studies the crystalline compounds present in nopal (Opuntia ficus-indica) cladodes. The identification of the crystalline structures was performed using X-ray diffraction, scanning electron microscopy, mass spectrometry, and Fourier transform infrared spectroscopy. The crystalline structures identified were calcium carbonate (calcite) [CaCO₃], calcium-magnesium bicarbonate [CaMg(CO₃)₂], magnesium oxide [MgO], calcium oxalate monohydrate [Ca(C₂O₄)•(H₂O)], potassium peroxydiphosphate [K₄P₂O₈] and potassium chloride [KCl]. The SEM images indicate that calcite crystals grow to dipyramidal, octahedral-like, prismatic, and flower-like structures; meanwhile, calcium-magnesium bicarbonate structures show rhombohedral exfoliation and calcium oxalate monohydrate is present in a drusenoid morphology. These calcium carbonate compounds have a great importance for humans because their bioavailability. This is the first report about the identification and structural analysis of calcium carbonate and calcium-magnesium bicarbonate in nopal cladodes, as well as the presence of magnesium oxide, potassium peroxydiphosphate and potassium chloride in these plants. The significance of the study of the inorganic components of these cactus plants is related with the increasing interest in the potential use of Opuntia as a raw material of products for the food, pharmaceutical, and cosmetic industries.     

Effect of polysaccharides from Opuntia ficus-indica (L.) cladodes on the healing of dermal wounds in the rat.

In traditional medicine extracts of polysaccharide-containing plants are widely employed for the treatment of skin and epithelium wounds and of mucous membrane irritation. The extracts of Opuntia ficus-indica cladodes are used in folk medicine for their antiulcer and wound-healing activities. The present study describes the wound-healing potential of two lyophilized polysaccharide extracts obtained from O. ficus-indica (L.) cladodes applied on large full-thickness wounds in the rat. When topically applied for 6 days, polysaccharides with a molecular weight (MW)>10(4)Da from O. ficus-indica cladodes induce a beneficial effect on cutaneous repair in this experimental model; in particular the topical application of O. ficus-indica extracts on skin lesions accelerates the reepithelization and remodelling phases, also by affecting cell-matrix interactions and by modulating laminin deposition. Furthermore, the wound-healing effect is more marked for polysaccharides with a MW ranging 10(4)-10(6)Da than for those with MW>10(6)Da, leading us to suppose that the fine structure of these polysaccharides and thus their particular hygroscopic, rheologic and viscoelastic properties may be essential for the wound-healing promoter activity observed.     

Changes in Osmotic Pressure and Mucilage during Low-Temperature Acclimation of Opuntia ficus-indica

Opuntia ficus-indica, a Crassulacean acid metabolism plant cultivated for its fruits and cladodes, was used to examine chemical and physiological events accompanying low-temperature acclimation. Changes in osmotic pressure, water content, low molecular weight solutes, and extracellular mucilage were monitored in the photosynthetic chlorenchyma and the water-storage parenchyma when plants maintained at day/night air temperatures of 30/20°C were shifted to 10/0°C. An increase in osmotic pressure of 0.13 megapascal occurred after 13 days at 10/0°C. Synthesis of glucose, fructose, and glycerol accounted for most of the observed increase in osmotic pressure during the low-temperature acclimation. Extracellular mucilage and the relative apoplastic water content increased by 24 and 10%, respectively, during exposure to low temperatures. These increases apparently favor the extracellular nucleation of ice closer to the equilibrium freezing temperature for plants at 10/0°C, which could make the cellular dehydration more gradual and less damaging. Nuclear magnetic resonance studies helped elucidate the cellular processes during ice formation, such as those revealed by changes in the relaxation times of two water fractions in the chlorenchyma. The latter results suggested a restricted mobility of intracellular water and an increased mobility of extracellular water for plants at 10/0°C compared with those at 30/20°C. Increased mobility of extracellular water could facilitate extracellular ice growth and thus delay the potentially lethal intracellular freezing during low-temperature acclimation.     

Severe Erwinia-Caused Damage on Opuntia ficus-indica in Italy

Erwinia carotovora subsp. carotovora. was consistently isolated from diseased tissues of prickly pear cactus ( Opuntia ficus-indica ) in Sicily. The bacterium was identified on the basis of morphological, cultural, biochemical and physiological characters as well as on the basis of pathogenicity. Symptoms were observed on cladophylls and fruit: watersoaked spots became brown and coalesced; the external tissue became dry and frequently cracked; the internal tissue turned brown to almost black. The disease was first localized in a few plantations, then seriously spread to all the modern intensive cultivations of prickly pear cacti.     

Arabinan-cellulose composite in Opuntia ficus-indica prickly pear spines

The ultrastructure of the spines decorating the cladodes of the cactus Opuntia ficus-indica was investigated by optical microscopy, scanning and transmission electron microscopy, wide angle X-ray, and solid state 13C NMR analyses. Each spine consisted of a compact parallel arrangement of slender cellulosic fibers (0.4 mm in length and 6-10 microm in diameter) with small lumens. The fibers were disencrusted by alkali and sodium chlorite bleaching, yielding a remarkable arabinan-cellulose (1:1) product. X-ray fiber diagrams of the spines before and after purification confirmed the presence of crystalline cellulose domains with molecular axis parallel to the spine axis. CP-MAS 13C T1 NMR data showed a strong interaction at a nanometric level of a fraction of the arabinan and the cellulose crystalline domains. By sequential hydrothermal extractions, followed by a trifluoroacetic acid treatment, a relatively pure cellulose was isolated while the extracted fibers became fibrillated into slender microfibrils having no more than 4-6 nm diameter. The hydrothermal extract yielded the alpha-L-arabinofuranan consisting of a chain of (1-->5)-linked L-arabinosyl residues with branching either at C-2 or C-3 or at both C-2 and C-3. Taken together, these observations suggest that the bulk of the spine fibers consists of an intimate composite of cellulose microfibrils embedded in an arabinan matrix.     

Physiological responses of Opuntia ficus-indica to growth temperature

The influences of various day/night air temperatures on net CO₂ uptake and nocturnal acid accumulation were determined for Opuntia ficus-indica, complementing previous studies on the water relations and responses to photosynthetically active radiation (PAR) for this widely cultivated cactus. As for other Crassulacean acid metabolism (CAM) plants, net nocturnal CO₂ uptake had a relatively low optimal temperature, ranging from 11°C for plants grown at day/night air temperatures of 10°C/0°C to 23°C at 45°C/35°C. Stomatal opening, which occurred essentially only at night and was measured by changes in water vapor conductance, progressively decreased as the measurement temperature was raised. The CO₂ residual conductance, which describes chlorenchyma properties, had a temperature optimum a few degrees higher than the optimum for net CO₂ uptake at all growth temperatures. Nocturnal CO₂ uptake and acid accumulation summed over the whole night were maximal for growth temperatures near 25°C/15°C, CO₂ uptake decreasing more rapidly than acid accumulation as the growth temperature was raised. At day/night air temperatures that led to substantial nocturnal acid accumulation (25°C/15°C.). 90% saturation of acid accumulation required a higher total daily PAR than at non-optimal growth temperatures (10°C/0°C and 35°C/25°C). Also, the optimal temperature of net CO₂ uptake shifted downward when the plants were under drought conditions at all three growth temperatures tested, possibly reflecting an increased fractional importance of respiration at the higher temperatures during drought. Thus, water status, ambient PAR, and growth temperatures must all be considered when predicting the temperature response of gas exchange for O. ficus-indica and presumably for other CAM plants.     

Carbon and water relations for developing fruits of Opuntia ficus-indica (L.) Miller, including effects of drought and gibberellic acid

Growth, gas exchange rates, and carbohydrate content were studied for developing fruits of the cultivated cactus Opuntia ficus-indica (L.) Miller, including effects of drought and exogenous gibberellic acid (GA3). Fruit development required 110 d from the time of bud differentiation to ripening at 80 d after anthesis, when the fruit mass averaged 67 g. Stomatal conductance and net CO2 uptake rates for fruits were higher during the night; they were maximal at 7 d before anthesis and decreased as development progressed. Fruits undergoing drought, imposed by detaching terminal stems bearing fruits, were 50% smaller than the control at 80 d after anthesis and did not ripen. Fruits injected with 2 ml of 500 ppm GA3 were 30% smaller than the control at 80 d after anthesis; they contained a large proportion of aborted seeds that produced a weak sink signal for dry mass accumulation. Gas exchange was higher at 21 d after anthesis for fruits treated with GA3. Total soluble sugars represented 40% of the fruit's dry mass until 45 d after anthesis, when the sugar content rapidly increased, reaching 90% at 73 d after anthesis. Such an increase was not observed for fruits treated with GA3, and the sugar content for fruits undergoing drought remained low throughout development. Starch content increased for developing fruits of O. ficus-indica until 14 d after anthesis and, except for the fruits undergoing drought, decreased thereafter. Fruit development for O. ficus-indica is apparently regulated by water availability as well as hormonal signals originating both within and outside the fruit.     

Aetiology of Opuntia ficus-indica malformations and stunting disease

The phytoplasma aetiology of epidemic stunting of cladodes and stunted growth observed in a cactus pear plantation in Carlentini (Sicily, Italy) was investigated with graft inoculation trials and PCR/restriction fragment length polymorphism analyses of 16S ribosomal RNA (rRNA) gene and S3 ribosomal protein gene. After sequencing of the 1525‐bp from 16S rRNA gene of both naturally infected and graft‐inoculated symptomatic cactus pear samples, phytoplasma TS belonging to ribosomal subgroup 16SrII‐C were identified as aetiological agents of this worldwide spread disease.     

Salt tolerance of prickly pear cactus (Opuntia ficus-indica)

In view of the need to exploit saline water resources in agriculture in arid zones, we investigated the salt tolerance of Opuntia ficus-indica in plants growing in solution culture. Salt (NaCl) was added in concentrations ranging from 5 (control) to 200 mol m^-3. Cladode growth was sensitive to salinity, being 60% of the control at 50 mol m^-3 NaCl. The root-to-stem ratio decreased significantly only at 200 mol m^-3. Various other parameters were studied, such as water content, Na, K and Cl content, osmotic pressure, and CO₂ uptake. Of these parameters the decreases in cladode water content and CO₂ uptake were related to the decrease in cladode growth. Raised salinity increased cladode osmotic pressure, which was associated with tissue dehydration. We concluded that osmotic adjustment does not occur in prickly pear under salt stress.