Post‐translational regulation of acid invertase activity by vacuolar invertase inhibitor affects resistance to cold‐induced sweetening of potato tubers

Cold‐induced sweetening (CIS) is a serious post‐harvest problem for potato tubers, which need to be stored cold to prevent sprouting and pathogenesis in order to maintain supply throughout the year. During storage at cold temperatures (below 10 °C), many cultivars accumulate free reducing sugars derived from a breakdown of starch to sucrose that is ultimately cleaved by acid invertase to produce glucose and fructose. When affected tubers are processed by frying or roasting, these reducing sugars react with free asparagine by the Maillard reaction, resulting in unacceptably dark‐coloured and bitter‐tasting product and generating the probable carcinogen acrylamide as a by‐product. We have previously identified a vacuolar invertase inhibitor (INH2) whose expression correlates both with low acid invertase activity and with resistance to CIS. Here we show that, during cold storage, overexpression of the INH2 vacuolar invertase inhibitor gene in CIS‐susceptible potato tubers reduced acid invertase activity, the accumulation of reducing sugars and the generation of acrylamide in subsequent fry tests. Conversely, suppression of vacuolar invertase inhibitor expression in a CIS‐resistant line increased susceptibility to CIS. The results show that post‐translational regulation of acid invertase by the vacuolar invertase inhibitor is an important component of resistance to CIS.     

Evidence for the complex relationship between free amino acid and sugar concentrations and acrylamide‐forming potential in potato

Free amino acids and reducing sugars participate in the Maillard reaction during high‐temperature cooking and processing. This results not only in the formation of colour, aroma and flavour compounds, but also undesirable contaminants, including acrylamide, which forms when the amino acid that participates in the reaction is asparagine. In this study, tubers of 13 varieties of potato (Solanum tuberosum), which had been produced in a field trial in 2010 and sampled immediately after harvest or after storage for 6 months, were analysed to show the relationship between the concentrations of free asparagine, other free amino acids, sugars and acrylamide‐forming potential. The varieties comprised five that are normally used for crisping, seven that are used for French fry production and one that is used for boiling. Acrylamide formation was measured in heated flour, and correlated with glucose and fructose concentration. In French fry varieties, which contain higher concentrations of sugars, acrylamide formation also correlated with free asparagine concentration, demonstrating the complex relationship between precursor concentration and acrylamide‐forming potential in potato. Storage of the potatoes for 6 months at 9°C had a significant, variety‐dependent impact on sugar and amino acid concentrations and acrylamide‐forming potential.     

Yield and cooking qualities of somaclonal variants of cv. Russet Burbank selected for resistance to common scab disease of potato

We previously obtained somaclonal variants of the important French fry processing cultivar Russet Burbank with significantly enhanced resistance to common scab disease. In this study we have shown the commercial merit of a proportion of these variants through comparison of relative yield and tuber quality with the parent cultivar Russet Burbank. Whilst we showed a weak negative correlation between tuber yield (as assessed by weight of tubers per plant) and relative disease resistance within selected variants, we identified several with equivalent yields to the parent cultivar. Furthermore, two disease-resistant variants (TC-RB8 and NZ-24B) consistently yielded more tuber mass than the parent. The majority of our Russet Burbank variants showed equivalent tuber quality characteristics (occurrence of defects, tuber specific gravity and dry matter content, and flesh colour) and cooking qualities (fry colour and presence of dark end defects) to the parent cultivar. Independent testing by a commercial French fry processor confirmed these quality characteristics. We present data demonstrating that highly common scab disease-resistant somaclonal variants of Russet Burbank have commercially acceptable tuber yield and quality characteristics, comparable to the industry standard and parent Russet Burbank cultivar. We also demonstrate the value of in vitro cell selection techniques for potato cultivar improvement.     

DNA variation at the invertase locus invGE/GF is associated with tuber quality traits in populations of potato breeding clones

Starch and sugar content of potato tubers are quantitative traits, which are models for the candidate gene approach for identifying the molecular basis of quantitative trait loci (QTL) in noninbred plants. Starch and sugar content are also important for the quality of processed products such as potato chips and French fries. A high content of the reducing sugars glucose and fructose results in inferior chip quality. Tuber starch content affects nutritional quality. Functional and genetic models suggest that genes encoding invertases control, among other things, tuber sugar content. The invGE/GF locus on potato chromosome IX consists of duplicated invertase genes invGE and invGF and colocalizes with cold-sweetening QTL Sug9. DNA variation at invGE/GF was analyzed in 188 tetraploid potato cultivars, which have been assessed for chip quality and tuber starch content. Two closely correlated invertase alleles, invGE-f and invGF-d, were associated with better chip quality in three breeding populations. Allele invGF-b was associated with lower tuber starch content. The potato invertase gene invGE is orthologous to the tomato invertase gene Lin5, which is causal for the fruit-sugar-yield QTL Brix9-2-5, suggesting that natural variation of sugar yield in tomato fruits and sugar content of potato tubers is controlled by functional variants of orthologous invertase genes.     

In-season heat stress compromises postharvest quality and low-temperature sweetening resistance in potato (Solanum tuberosum L.)

Key message

High soil temperature during bulking and maturation of potatoes alters postharvest carbohydrate metabolism to attenuate genotypic resistance to cold-induced sweetening and accelerate loss of process quality.

Abstract

The effects of soil temperature during tuber development on physiological processes affecting retention of postharvest quality in low-temperature sweetening (LTS) resistant and susceptible potato cultivars were investigated. ‘Premier Russet’ (LTS resistant), AO02183-2 (LTS resistant) and ‘Ranger Russet’ (LTS susceptible) tubers were grown at 16 (ambient), 23 and 29 °C during bulking (111–164 DAP) and maturation (151–180 DAP). Bulking at 29 °C virtually eliminated yield despite vigorous vine growth. Tuber specific gravity decreased as soil temperature increased during bulking, but was not affected by temperature during maturation. Bulking at 23 °C and maturation at 29 °C induced higher reducing sugar levels in the proximal (basal) ends of tubers, resulting in non-uniform fry color at harvest, and abolished the LTS-resistant phenotype of ‘Premier Russet’ tubers. AO02183-2 tubers were more tolerant of heat for retention of LTS resistance. Higher bulking and maturation temperatures also accelerated LTS and loss of process quality of ‘Ranger Russet’ tubers, consistent with increased invertase and lower invertase inhibitor activities. During LTS, tuber respiration fell rapidly to a minimum as temperature decreased from 9 to 4 °C, followed by an increase to a maximum as tubers acclimated to 4 °C; respiration then declined over the remaining storage period. The magnitude of this cold-induced acclimation response correlated directly with the extent of buildup in sugars over the 24-day LTS period and thus reflected the effects of in-season heat stress on propensity of tubers to sweeten and lose process quality at 4 °C. While morphologically indistinguishable from control tubers, tubers grown at elevated temperature had different basal metabolic (respiration) rates at harvest and during cold acclimation, reduced dormancy during storage, greater increases in sucrose and reducing sugars and associated loss of process quality during LTS, and reduced ability to improve process quality through reconditioning. Breeding for retention of postharvest quality and LTS resistance should consider strategies for incorporating more robust tolerance to in-season heat stress.     

Acrylamide in processed potato products: progress made and present status

Concerns related to higher levels of acrylamide in processed carbohydrate-rich foods, especially in fried potato products, are well known. This article provides updates on various aspects of acrylamide in processed potato products including mechanisms of acrylamide formation and health risks due to its intake. Levels of reducing sugars in potatoes are considered as a main factor contributing towards the formation of acrylamide in processed potato products. Useful approaches in lowering the levels of reducing sugars such as use of suitable varieties, storage methods, storage temperature and duration of storage are described and discussed. Importance and practical utility of various steps before and during the processing that can contribute in reducing the final concentration of acrylamide are highlighted. Progress made and present status of potato processing industry in India are part of this article. The article describes varietal improvement and spread of short-term and long-term storage technologies in India and their contribution towards round the year availability of processing-grade potatoes to the processing industries and how all this has helped in achieving reduced levels of acrylamide in chips and French fries. Outcome and implications of cold-induced sweetening tolerance in potatoes are presented along with other management practices and strategies that can lower the acrylamide levels in processed potato products. Future lines of work have been suggested to make the consumption of fried potato products safer.     

Photosynthetic assimilation of 14C into amino acids in potato (Solanum tuberosum) and asparagine in the tubers

Asparagine is the predominant free amino acid in potato tubers and the present study aimed to establish whether it is imported from the leaves or synthesised in situ. Free amino acid concentrations are important quality determinants for potato tubers because they react with reducing sugars at high temperatures in the Maillard reaction. This reaction produces melanoidin pigments and a host of aroma and flavour volatiles, but if free asparagine participates in the final stages, it results in the production of acrylamide, an undesirable contaminant. ¹⁴CO₂ was supplied to a leaf or leaves of potato plants (cv. Saturna) in the light and radioactivity incorporated into amino acids was determined in the leaves, stems, stolons and tubers. Radioactivity was found in free amino acids, including asparagine, in all tissues, but the amount incorporated in asparagine transported to the tubers and stolons was much less than that in glutamate, glutamine, serine and alanine. The study showed that free asparagine does not play an important role in the transport of nitrogen from leaf to tuber in potato, and that the high concentrations of free asparagine that accumulate in potato tubers arise from synthesis in situ. This indicates that genetic interventions to reduce free asparagine concentration in potato tubers will have to target asparagine metabolism in the tuber.     

Engineering and biotechnological aspects for the manufacturing of high quality fried potato products

Fried potato products have become very popular foods over the last decades. High quality standards have been established for these products by the food industry including uniform brown color and crispness. During frying, Maillard reactions takes place which contribute to color and taste development in these products. However, safety aspects are also influenced by these reactions, e.g., acrylamide formation. Maintaining high safety standards as well as the expected quality requires systematic research based on an integrated approach including all relevant variables, e.g., raw material properties, processing conditions and equipment concepts. Selected results of these investigations are presented and discussed, regarding influence of composition, e.g., precursor levels for Maillard reactions, treatment of raw materials and addition of reactants to frying fat. It has been demonstrated that a combined treatment of the potato sticks by coating of product surfaces and partial pre-drying can be successfully applied to produce well-browned French fries with lower acrylamide contents. Reductions up to 75% could be reached compared to samples without treatment. Furthermore, addition of a water/oil emulsion containing glutamine in the aqueous phase has been shown to influence Maillard reactions at the product surface, resulting in lower acrylamide contents at the same state of browning.     

Inhibition of Potato Tuber Invertase by an Endogenous Inhibitor: EFFECTS OF SALTS, pH, TEMPERATURE, AND SUGARS ON BINDING

Binding between potato tuber invertase and its endogenous inhibitor followed second-order reaction kinetics. Binding rates were diminished by the presence of various inorganic salts, MgCl₂ being especially effective. This effect of MgCl₂ was used in binding rate studies by adding the salt with sucrose to reduce binding during assay of previously unbound activity. The optimal pH for binding was about 4.8, similar to the optimal pH for catalytic activity of invertase. The optimal temperature for binding was about 45 C, approximately 5 C less than the optimum for catalytic activity. Sucrose at concentrations as low as 2 millimolar slowed binding; reducing sugars had little or no effect on binding or on catalytic activity.     

Soluble acid invertase determines the hexose-to-sucrose ratio in cold-stored potato tubers

Cold storage of potato (Solanum tuberosum L.) tubers is known to cause accumulation of reducing sugars. Hexose accumulation has been shown to be cultivar-dependent and proposed to be the result of sucrose hydrolysis via invertase. To study whether hexose accumulation is indeed related to the amount of invertase activities, two different approaches were used: (i) neutral and acidic invertase activities as well as soluble sugars were measured in cold-stored tubers of 24 potato cultivars differing in the cold-induced accumulation of reducing sugars and (ii) antisense potato plants with reduced soluble acid invertase activities were created and the soluble sugar accumulation in cold-stored tubers was studied. The cold-induced hexose accumulation in tubers from the different potato cultivars varied strongly (up to eightfold). Large differences were also detected with respect to soluble acid (50-fold) and neutral (5-fold) invertase activities among the different cultivars. Although there was almost no correlation between the total amount of invertase activity and the accumulation of reducing sugars there was a striking correlation between the hexose/sucrose ratio and the extractable soluble invertase activitiy. To exclude the possibility that other cultivar-specific features could account for the obtained results, the antisense approach was used to decrease the amount of soluble acid invertase activity in a uniform genetic background. To this end the cDNA of a cold-inducible soluble acid invertase (EMBL nucleicacid database accession no. X70368) was cloned from the cultivar Desirée, and transgenic potato plants were created expressing this cDNA in the antisense orientation under control of the constitutive 35S cauliflower mosaic virus promotor. Analysis of the harvested and cold-stored tubers showed that inhibition of the soluble acid invertase activity leads to a decreased hexose and an increased sucrose content compared with controls. As was already found for the different potato cultivars the hexose/sucrose ratio decreased with decreasing invertase activities but the total amount of soluble sugars did not significantly change. From these data we conclude that invertases do not control the total amount of soluble sugars in coldstored potato tubers but are involved in the regulation of the ratio of hexose to sucrose.The authors are grateful to Heike Deppner and Christiane Prüßner for tuber harvest and technical assistance during the further analysis. We thank Andrea Knospe for taking care of tissue culture, Birgit Schäfer for patient photographic work, Hellmuth Fromme and the greenhouse personnel for attending plant growth and development and Astrid Basner for elucidating the sequence of clone INV-19. The work was supported by the Bundesministerium für Forschung und Technologie (BMFT).     

Translucent tissue defect in potato (Solanum tuberosum L.) tubers is associated with oxidative stress accompanying an accelerated aging phenotype

Translucent tissue defect (TTD) is an undesirable postharvest disorder of potato tubers characterized by the development of random pockets of semi-transparent tissue containing high concentrations of reducing sugars. Translucent areas turn dark during frying due to the Maillard reaction. The newly released cultivar, Premier Russet, is highly resistant to low temperature sweetening, but susceptible to TTD. Symptoms appeared as early as 170 days after harvest and worsened with time in storage (4–9 °C, 95 % RH). In addition to higher concentrations of glucose, fructose and sucrose, TTD resulted in lower dry matter, higher specific activities of starch phosphorylase and glc-6-phosphate dehydrogenase, higher protease activity, loss of protein, and increased concentrations of free amino acids (esp. asparagine and glutamine). The mechanism of TTD is unknown; however, the disorder has similarities with the irreversible senescent sweetening that occurs in tubers during long-term storage, where much of the decline in quality is a consequence of progressive increases in oxidative stress with advancing age. The respiration rate of non-TTD ‘Premier Russet’ tubers was inherently higher (ca. 40 %) than that of ‘Russet Burbank’ tubers (a non-TTD cultivar). Moreover, translucent tissue from ‘Premier Russet’ tubers had a 1.9-fold higher respiration rate than the average of non-translucent tissue and tissue from non-TTD tubers. Peroxidation of membrane lipids during TTD development resulted in increased levels of malondialdehyde and likely contributed to a measurable increase in membrane permeability. Superoxide dismutase and catalase activities and the ratio of oxidized to total glutathione were substantially higher in translucent tissue. TTD tubers also contained twofold less ascorbate than non-TTD tubers. TTD appears to be a consequence of oxidative stress associated with accelerated aging of ‘Premier Russet’ tubers.     

Acrylamide in food: Progress in and prospects for genetic and agronomic solutions

Acrylamide is a processing contaminant and Group 2a carcinogen that was discovered in foodstuffs in 2002. Its presence in a range of popular foods has become one of the most difficult problems facing the food industry and its supply chain. Wheat, rye and potato products are major sources of dietary acrylamide, with biscuits, breakfast cereals, bread (particularly toasted), crispbread, batter, cakes, pies, French fries, crisps and snack products all affected. Here we briefly review the history of the issue, detection methods, the levels of acrylamide in popular foods and the risk that dietary acrylamide poses to human health. The pathways for acrylamide formation from free (non‐protein) asparagine are described, including the role of reducing sugars such as glucose, fructose and maltose and the Maillard reaction. The evolving regulatory situation in the European Union and elsewhere is discussed, noting that food businesses and their suppliers must plan to comply not only with current regulations but with possible future regulatory scenarios. The main focus of the review is on the genetic and agronomic approaches being developed to reduce the acrylamide‐forming potential of potatoes and cereals and these are described in detail, including variety selection, plant breeding, biotechnology and crop management. Obvious targets for genetic interventions include asparagine synthetase genes, and the asparagine synthetase gene families of different crop species are compared. Current knowledge on crop management best practice is described, including maintaining optimum storage conditions for potatoes and ensuring sulphur sufficiency and disease control for wheat.     

Inhibitory Effect of Maillard Reaction Products on Growth of the Aerobic Marine Hyperthermophilic Archaeon Aeropyrum pernix

It was found that the growth of Aeropyrum pernix was severely inhibited in a medium containing reducing sugars and tryptone due to the formation of Maillard reaction products. The rate of the Maillard browning reaction was markedly enhanced under aerobic conditions, and the addition of Maillard reaction products to the culture medium caused fatal growth inhibition.