Postharvest Physiological Deterioration Research Articles

Constituents and secondary metabolite natural products in fresh and deteriorated cassava roots

A phytochemical analysis of cassava (Manihot esculenta Crantz) fresh roots and roots suffering from post-harvest physiological deterioration (PPD) has been carried out. The first isolation and identification of galactosyl diacylglycerides from fresh cassava roots is reported, as well as beta-carotene, linamarin, and beta-sitosterol glucopyranoside. The hydroxycoumarin scopoletin and its glucoside scopolin were identified from cassava roots during PPD, as well as trace quantities of esculetin and its glucoside esculin. There is no isoscopoletin in cassava roots during PPD.

Investigation of Biosynthetic Pathways to Hydroxycoumarins During Post‐Harvest Physiological Deterioration in Cassava Roots by Using Stable Isotope Labelling

Cassava (Manihot esculenta Crantz) is an important starch-rich crop, but the storage roots only have a short shelf-life due to post-harvest physiological deterioration (PPD), which includes the over-production and polymerisation of hydroxycoumarins. Key aspects of coumarin secondary-metabolite biosynthesis remain unresolved. Here we exploit the accumulation of hydroxycoumarins to test alternative pathways for their biosynthesis. Using isotopically labelled intermediates (p-coumarate-2-(13)C, caffeate-2-(13)C, ferulate-2-(13)C, umbelliferone-2-(18)O and esculetin-2-(18)O), we show that the major biosynthetic pathway to scopoletin and its glucoside, scopolin, in cassava roots during PPD is through p-coumaric, caffeic and then ferulic acids. An alternate pathway through 2',4'-dihydroxycinnamate and umbelliferone leads to esculetin and esculin. We have used C(18)O(2)-carboxylate-labelled cinnamic and ferulic acids, and feeding experiments under an atmosphere of (18)O(2), to investigate the o-hydroxylation and cyclisation steps. We demonstrate that the major pathway is through o-hydroxylation and not via a proposed spirolactone-dienone intermediate.

Biosynthesis of scopoletin and scopolin in cassava roots during post-harvest physiological deterioration: The E- Z-isomerisation stage

Two to three days after harvesting, cassava ( Manihot esculenta Crantz) roots suffer from post-harvest physiological deterioration (PPD) when secondary metabolites are accumulated. Amongst these are hydroxycoumarins (e.g. scopoletin and its glucoside scopolin) which play roles in plant defence and have pharmacological activities. Some steps in the biosynthesis of these molecules are still unknown in cassava and in other plants. We exploit the accumulation of these coumarins during PPD to investigate the E-Z-isomerisation step in their biosynthesis. Feeding cubed cassava roots with E-cinnamic-3,2′,3′,4′,5′,6′- d 5 acid gave scopoletin- d 2. However, feeding with E-cinnamic-3,2′,3′,4′,5′,6′- d 6 and E-cinnamic-2,3,2′,3′,4′,5′,6′- d 7 acids, both gave scopoletin- d 3, the latter not affording the expected scopoletin- d 4. We therefore synthesised and fed with E-cinnamic-2- d 1 when unlabelled scopoletin was biosynthesised. Solely the hydrogen (or deuterium) at C2 of cinnamic acid is exchanged in the biosynthesis of hydroxycoumarins. If the mechanism of E-Z-cinnamic acid isomerisation were photochemical, we would not expect to see the loss of deuterium which we observed. Therefore, a possible mechanism is an enzyme catalysed 1,4-Michael addition, followed by σ-bond rotation and hydrogen (or deuterium) elimination to yield the Z-isomer. Feeding the roots under light and dark conditions with E-cinnamic-2,3,2′,3′,4′,5′,6′- d 7 acid gave scopoletin- d 3 with no significant difference in the yields. We conclude that the E-Z-isomerisation stage in the biosynthesis of scopoletin and scopolin, in cassava roots during PPD, is not photochemical, but could be catalysed by an isomerase which is independent of light.

Sequencing analysis of 20,000 full-length cDNA clones from cassava reveals lineage specific expansions in gene families related to stress response

BackgroundCassava, an allotetraploid known for its remarkable tolerance to abiotic stresses is an important source of energy for humans and animals and a raw material for many industrial processes. A full-length cDNA library of cassava plants under normal, heat, drought, aluminum and post harvest physiological deterioration conditions was built; 19968 clones were sequence-characterized using expressed sequence tags (ESTs).ResultsThe ESTs were assembled into 6355 contigs and 9026 singletons that were further grouped into 10577 scaffolds; we found 4621 new cassava sequences and 1521 sequences with no significant similarity to plant protein databases. Transcripts of 7796 distinct genes were captured and we were able to assign a functional classification to 78% of them while finding more than half of the enzymes annotated in metabolic pathways in Arabidopsis. The annotation of sequences that were not paired to transcripts of other species included many stress-related functional categories showing that our library is enriched with stress-induced genes. Finally, we detected 230 putative gene duplications that include key enzymes in reactive oxygen species signaling pathways and could play a role in cassava stress response features.ConclusionThe cassava full-length cDNA library here presented contains transcripts of genes involved in stress response as well as genes important for different areas of cassava research. This library will be an important resource for gene discovery, characterization and cloning; in the near future it will aid the annotation of the cassava genome.

Towards identifying the full set of genes expressed during cassava post-harvest physiological deterioration

Storage roots of cassava (Manihot esculenta Crantz) exhibit a rapid post-harvest physiological deterioration (PPD) response that can occur within 24-72 h of harvest. PPD is an enzymatically mediated oxidative process with parallels to plant wound, senescence and defence responses. To characterise those genes that show significant change in expression during the PPD response we have used cDNA microarray technology to carry out a large-scale analysis of the cassava root transcriptome during the post-harvest period. We identified 72 non-redundant expressed sequence tags which showed altered regulation during the post-harvest period. Of these 63 were induced, whilst 9 were down-regulated. RNA blot analysis of selected genes was used to verify and extend the microarray data. Additional microarray hybridisation experiments allowed the identification of 21 root-specific and 24 root-wounding-specific sequences. Many of the up-regulated and PPD-specific expressed sequence tags were predicted to play a role in cellular processes including reactive oxygen species turnover, cell wall repair, programmed cell death, ion, water or metabolite transport, signal transduction or perception, stress response, metabolism and biosynthesis, and activation of protein synthesis.

Oxidative stress responses during cassava post-harvest physiological deterioration

A major constraint to the development of cassava (Manihot esculenta Crantz) as a crop to both farmers and processors is its starchy storage roots' rapid post-harvest deterioration, which can render it unpalatable and un-marketable within 24-72 h. An oxidative burst occurs within 15 min of the root being injured, that is followed by the altered regulation of genes, notably for catalase and peroxidase, related to the modulation of reactive oxygen species, and the accumulation of secondary metabolites, some of which show antioxidant properties. The interactions between these enzymes and compounds, in particular peroxidase and the coumarin, scopoletin, are largely confined to the vascular tissues where the visible symptoms of deterioration are observed. These, together with other data, are used to develop a tentative model of some of the principal events involved in the deterioration process.

Oxidative stress responses during cassava post-harvest physiological deterioration

Oxidative stress responses during cassava post-harvest physiological deterioration

Lipid peroxidation, activities of superoxide dismutase and catalase during post-harvest deterioration of cassava (Manihot esculenta Crantz) root tubers

The alterations in the level of lipid peroxidation, activities of superoxide dismutase (SOD) and catalase were studied during physiological post-harvest deterioration of cassava (Manihot esculenta Crantz) root tubers for four days. The peak level of lipid peroxidation was observed on the 1st day, which also coincided with the peak activities of total SOD, CuZnSOD and catalase. With the exception of the 1st day, the activity of Mn-SOD was consistently higher than that of CuZnSOD. The data presented here may suggest that the enhanced level of lipid peroxidation on the 1st day and the sustained decrease in the activities of SOD and catalase could contribute to the post-harvest deterioration of cassava root tubers.

Mapping wound-response genes involved in post-harvest physiological deterioration (PPD) of cassava (Manihot esculenta Crantz)

The genome locations of the wound-response genes that were expressedduring the post-harvest physiological deterioration (PPD) of cassava, suchas phenylalanine ammonia lyase, β-1.3 glucanase, hydroxyprolinerich glycoprotein, catalase, 1-aminocyclopropane 1-carboxylate, cysteineprotease inhibitor, aspartic protease, a partial cDNA for serine/threonineprotein kinase and peroxidase, have been identified on the frameworkmolecular genetic map of cassava. Also, molecular markers linked toputative quantitative trait loci (QTLs) influencing PPD of cassava weremapped using an F1mapping population derived from elite parentallines (TMS 30572 × cm 2177-2). A molecular linkage map previouslyconstructed based on the segregation of 240 RFLP, 100 RAPD, 85microsatellite and five isoenzyme markers on 144 F1 individuals wasused for the QTL mapping.A set of 10 molecular markers with a significant association with putativeQTLs for PPD were identified based on probability values < 0.005in order to minimize the detection of false positives. Based on single-markerregression, eight putative QTLs located on the linkage groups G, P, L, U,and X of the female-derived framework map were found to explain between 5–12% of the phenotypic variance of the PPD. In the male-derived frameworkmap, two putative QTLs on linkage groups C and L explained 13% and11% of this variance, respectively. This study thus identified the majorgenome regions of cassava related to physiological post-harvestdeterioration, thereby providing tools for the identification of gene(s)controlling this trait.

Isolation and characterisation of a cassava catalase expressed during post-harvest physiological deterioration

Storage roots of cassava undergo a rapid, endogenous, post-harvest deterioration response that is thought to involve oxidative processes. A cassava catalase (MecCAT1) was isolated from a root cDNA library. The transcript is expressed predominantly in roots with little expression in leaves. Catalase enzyme activity and MecCAT1 transcript expression during the post-harvest period were compared in highly susceptible and less susceptible cultivars and suggest that high levels of catalase activity may play a role in delaying the deterioration response.

Molecular analysis of differentially expressed genes during postharvest deterioration in cassava (Manihot esculenta Crantz) tuberous roots

One of the major problems for cassava is the rapid deterioration after harvesting cassava tuberous roots, which limits the possibilities for production and distribution of cassava in the world. Postharvest deterioration is an inherent problem for cassava since wounding and mechanical damage of the tuberous roots cannot be prevented during harvesting, which includes postharvest physiological deterioration (PPD) and secondary deterioration. To date, the molecular mechanism and biochemical pathways of PPD are poorly understood. The aim of this project, which is focusing on the early stages (first 72 hrs), is to gain molecular insight and identify important metabolic pathways during the process of PPD in cassava tuberous roots. Finally by reverse genetic approaches to delay or even prevent the process of PPD in cassava tuberous roots. By using a new RNA fingerprinting method, called cDNA-AFLP, we have screened more than 6,000 TDFs (Transcript Derived Fragments) via up to 100 primer combinations during the early process of PPD in cassava. Only 10% of the TDFs are developmentally regulated, while the other90% are expressed throughout the process of PPD in cassava tuberous roots. Furthermore, in order to set up a functional catalogue of differentially expressed genes during PPD, 70 TDFs were selected and isolated based on their expression patterns, which were either up-regulated,down-regulated or transiently induced. Around 40 of these TDFs were found to be similar with known genes in databases. The other 30 TDFs were present mostly genes without known function. Through data analysis,it is shown that important biochemical and physiological processes, such as notably oxygen stress, carbohydrate metabolism, protein metabolism and phenolic compounds synthesis, are involved in PPD in cassava tuberous roots.

Hydrogen peroxide and flavan-3-ols in storage roots of cassava (Manihot esculenta crantz) during postharvest deterioration.

Cassava storage roots are an important staple food throughout the lowland humid tropics. However, cassava suffers from a poorly understood storage disorder, known as postharvest physiological deterioration (PPD), which constrains its exploitation. In an attempt to broaden the understanding of PPD, nine different cassava cultivars were analyzed for specific compounds accumulating during the process. The production of hydrogen peroxide (H(2)O(2)) is involved in the early stages of PPD in cassava roots. H(2)O(2) was quantified and localized histochemically at the tissue and cell level in deteriorating roots. This reactive oxygen species accumulated during the first 24 h after harvest, especially in the inner parenchymatic tissue. Three flavan-3-ols, (+)-catechin, (+)-catechin gallate, and (+)-gallocatechin, accumulated during the storage of cassava roots. However, these potential antioxidants cannot be related to early storage disorders or wound responses because they start to accumulate only after 4-6 days.

The effect of pre-harvest pruning of cassava upon root deterioration and quality characteristics

One of the major constraints of cassava as a crop is its perishability. Physiological deterioration, parenchymal blue-black vascular streaking, often starts within 24 h after harvest. This paper presents the results of a detailed study of the effects of pre-harvest pruning upon post-harvest physiological deterioration (PPD) and some other quality characteristics. Six cultivars, grown at CIAT (Centro International de Agricultura Tropical), with varying intrinsic susceptibility to PPD), were assessed at pruning-harvest intervals of 0, 2, 4, 6, 8, 10, 15, 20, 25, 28 and 39 days. After harvesting, the roots were analysed. For the unpruned plants a low susceptibility was found to coincide with a low dry matter content and a high sugar content. After pruning, the susceptibility for all cultivars was drastically reduced, reaching a minimum of around 25% of the original value for a pruning-harvest interval of up to 25 days. Beyond this interval the plants slowly develop new leaf canopy, normal assimilation sets in again and the starch content increases. Analysis of the cassava roots revealed a relationship between the combined sugar and starch contents and the interval duration, and that sugar and starch contents were inversely related to each other. The sugar content increased with the interval period, probably as a result of starch hydrolysis. Other properties such as the contents of dry matter, cyanogen, scopoletin, amylose and reducing sugars and the starch pasting properties were not affected by pruning to a comparable, interval-dependent, extent. It is concluded that the sugar content, ie the sugaristarch ratio, of cassava roots is positively related to their resistance to post-harvest physiological deterioration. (Resume d'auteur)

Iron, Carotene, and Ascorbic Acid in Cassava Roots and Leaves

The cassava core collection (601 genotypes) was evaluated for root and leaf contents of micronutrient minerals, ascorbic acid, and carotene. Wide genetic variability was observed for all measurements, indicating that there is good potential for exploiting and improving the nutritive value of cassava. There seems to be little correlation between the levels of any micronutrient in roots and leaves. There was no clear association between carotene and ascorbic acid concentrations. A genetic study of the progeny of a cross between yellow and white parents indicated control of the yellow trait by only two genes. The stability of vitamins after three commonly used processing procedures was evaluated in a sample of 26 genotypes. A higher proportion of the original vitamin content survived boiling, whereas solar drying resulted in the highest losses. Carotene was more stable than ascorbic acid. In a limited number of lines, there was some indication that higher vitamin content was associated with decreased post-harvest physiological deterioration. Since it is well established that β-carotene and ascorbic acid can enhance the absorption and internal transport of dietary iron and zinc from plant sources, yellow varieties of cassava have potential to address not only vitamin deficiencies per se, but also iron-deficiency anaemia and zinc deficiency. Further, the use of the leaves as a vegetable, as is done in several African countries, can complement the use of the root as a staple because of the high nutrient density of the leaves. The potential to improve the nutritive potential of cassava is exciting.