Our institute is beneficiary of the project of the 7. FP HighTech Europe, details see http://www.hightecheurope.eu/.

The results achieved in various projects, in research suported by grants and by the work under the research plan are presented in the form of research or technical papers, patents, or papers read at workshops and conferences.

Most important results achieved in 2009
authors from other organizations are marked by an asterisk (*)
 

Štumr*F., Gabrovská D., Rysová J., Hanák P., Plicka*J., Tomková*K., Cuhra*P., Kubík*M., Baršová*S., Karšulínová*L., Bulawová*H., Brychta*J.
ELISA kit for beta-lactoglobulin determination – Collaborative study.
J.AOAC. 2009; 92(5): 1519-1525, ISSN 0004-5756.
A collaborative study in 6 laboratories was performed to prove the validation of ELISA method developed for quantitative beta-lactoglobulin determination in foods. The ELISA kit used for this study is based on rabbit polyclonal antibody. This kit does not produce any false positive results or cross-reactivity with broad range of food matrix with zero content of milk proteins. In-house validation did not prove false positivity or cross-reactivity with broad range of food matrix with zero content of milk protein. All participants obtained the beta-lactoglobulin kit with standard operational procedure, the list of the samples, samples and a protocol for test results recording. The study included 14 food samples and 6 spiked samples: bread, 2 soybean desserts, butter, chicken ham, chicken meat, wheat flour, long grain rice, jelly, 2 whey drinks, crackers, bitter chocolate and 2 spiked samples of rice, 2 spiked samples of wheat flour and 2 spiked samples of chicken meat. Nine samples of food matrix with zero content of milk proteins showed beta-lactoglobulin content lower than the first standard (beta-lactoglobulin content 0.15 mg/kg). Two samples of food matrix with zero content of milk proteins revealed beta-lactoglobulin contents higher than standard 3 (1.5 mg BLG/100 g) and standard 4 (5.0 mg BLG/100g). Three food samples containing milk as ingredient were tested as positive and also all spiked samples were evaluated as positive. The statistical tests (Cochran, Dixon and Mandel) and analysis of variance (ANOVA) were used for the evaluation of collaborative study results. Repeatability and reproducibility limits as well as the limit of quantification (LOQ, 0.22 mg BLG/kg) and the limit of detection (LOD, 0.07 mg BLG/kg) for the kit were calculated.

Dostálek*P., Gabrovská D., Rysová J., Mena*MC., Hernando*A., Méndez*E., Chmelík*J., Šalplachta*J.
Determination of gluten in glucose syrups.
Journal of Food Composition and Analyses. 2009; 22: 762 -765, ISSN 0889-1575.
The gluten content in various glucose syrups was determined by two sandwich ELISA methods and one competitive ELISA method. Different extraction solutions were used for ELISA methods. MALDI-TOF mass spectrometry and SDS-PAGE were also carried out as a complementary technique to ELISA methods. The analysis proved that glucose syrups analyzed in this work and used in many common and gluten-free food products are safe for patients suffering from celiac disease. Four food products containing glucose syrup were analyzed with satisfactory results as well. One sample of chocolate bar contained gluten amounts below the current limit for gluten-free foods. Gluten content above this limit was found in this chocolate bar after cocktail-gelatine extraction. This product is not labelled as gluten-free food. This fact shows that the analyses of gluten-free products are not simple and require more work and attention.

Tomková*K., Štumr*F., Dvorská*P., Šafářová*P., Rysová J., Gabrovská D., Hanák P., Plicka*J.
Methods for the Determination of Allergenic Substances in Foods.
Czech J.Food Sci. 2009; 27(Special Issue): 369 – 371, ISSN 1212-1800.
Within the framework of the research project ELISA methods for the quantitative determination of allergenic substances in foodstuff and raw materials were developed. ELISA kits for allergenic proteins of milk (casein, beta-lactoglobulin and BSA), egg white proteins and mustard proteins were validated and collaborative studies were performed to prove the validation of the ELISA methods developed. Various methods of extraction were tested. The parameters like the limit of detection, limit of quantification, robustness, repeatability and accuracy were determined. A broad range of zero matrices for allergens were tested as well. The ELISA kits are suitable for the determination of allergens according to EU legislation Directive 2005/26/EC and Directive 2006/142/EC in the laboratories focused on this topic.

Ehrenbergerová*J., Březinová Belcredi*N., Kopáček*J., Melišová*L., Hrstková*P., Macuchová*A., Vaculová*K., Paulíčková I.
Antioxidant Enzymes in Barley Green Biomass.
Plant Foods Hum. Nutr. (2009) 64:122-128, ISSN 0921-9668 (print) ISSN 1573-9104 (online).
The first sampling of green matter of young barley plants (in growth phase DC 29) contained a statistically significant higher superoxide dimustase (SOD) and catalase (CAT) activity as well as higher concentration of vitamin C, in comparison with the later sampling (in DC 31). In the average of years and localities, in the 1st sampling the variety Sebastian and the line KM 1910 showed a significantly higher CAT activity (935 a 907 U. g^-1), compared to the variety Malz. Variety Sebastian had significantly higher CAT activity in the locality of Kroměříž (908 U. g^-1) and line KM 1910 in the locality of Žabčice (832 U. g^-1). Statistically significant higher SOD activity was found in variety Sebastian (574 U. g^-1) in the locality of Žabčice in the 1st sampling; the total average for the whole experiment (486 U. g^-1) was statistically significant higher if compared to the variety Malz and the line KM1910. Judging by the content of vitamin C, the most suitable were the locality of Kroměříž with the significantly higher average biomass activity (583 mg.100 g^-1), and the variety Sebastian with the average total-experiment activity 569 mg.100 g-1 in the 1st sampling, yet not differing significantly from the variety Malz and the line KM1910. Significant varietal influences on SOD activity and interactional influences were determined: those of varieties with biomass samplings, localities and growing years, as well as of localities with growing years. The 3rd sampling (in DC 31) was inappropriate for the said phytochemicals due to their statistically significant lower content and the high percentage of fibre. It can be stated that in the research period (2005-2007) the green biomass of young spring barley plants in growth phase DC 29 was an important source of vitamin C and of the enzymes catalase and superoxide dismutase. Because of the presence of these bioactive substances significant for health prevention, young green barley matter has a potential of being used primarily in the area of food supplements.

Prošková A., Kučera J., Kopicová Z.
Comparison of acid and alkali catalyzed transesterification of rendering plant fat with methanol.
Chem. Listy. 2009; 103(12): 1034–1036, ISSN 0009-2770.
Acid- and alkali- catalyzed transesterifications of fat from rendering plant as a source for production of biodiesel fuels were compared. The alkali-catalyzed reaction gives lower conversions of triglycerides, probably due to high contents of free fatty acids. The acid-catalyzed transesterification is more appropriate as it gives high yields of triglycerides but requires higher temperature and long reaction time.

Kronek*J., Žitný*R., Horný*L., Chlup*H., Beran M.
Mechanical properties of artery-artery connection based upon transglutaminase cross-linked gelatin.
Metallurgy. 2010; 49(2): 356-360, ISSN 0543-5846.
The possibility of surgical repairing of blood vessels by adhesive biocompatible glues represents an alternative to the conventional sewing techniques. The aim of our study was to evaluate mechanical properties of adhesively connected arteries by a cross linked gelatin. Series of quasistatic uniaxial tensile tests of two overlapping arterial strips bonded by a two-component glue (gelatin linked by the enzyme transglutaminase - TGA) was carried out, giving basic relationship between the loading forces and the sample extension, ultimate stresses etc. The 3D digital image correlation system (DIC) gave more details, local deformations, and thus the mutual slipping of the bonded strips could be evaluated (in this way the contributions of artery and the bond itself were separated). The effect of TGA and gelatin concentrations were estimated on the basis of observed data. Recorded maximum stresses were rather small (only tens of kPa), even if the subsequent histological analysis proved that the applied glue cross-linked a thin layer of the connected tissue too (at least partially).

Beran M., Klubal*R., Molik P., Strohalm J., Urban M., Klaudyova*A. A., Prajzlerova*K.
Influence of high hydrostatic pressure on tryptic and chymotryptic hydrolysis of cow milk proteins.
High Pressure Research 2009; 29(1): 23-27. Special Issue: Proceedings of the XLVIth European High Pressure Research Group (EHPRG 46) Meeting. 7-12 September 2008, Valencia, Spain. ISSN 0895-7959 (print), 1477-2299 (online).
This paper describes the effect of isostatic pressure 500 MPa on tryptic and chymotryptic hydrolysis of α- and β-casein, bovine serum albumin (BSA), β-lactoglobulin (β-Lg) and α- lactalbumin (α-La). Digestion was also conducted at atmospheric pressure. The extent of hydrolysis and peptide profiles were analysed by gel-permeation and reverse-phase high-performance liquid chromatographies. The residual immunochemical reactivities of the protein hydrolysates were assessed by the Streptavidin ImmunoCAP system (Phadia) for determination of specific immunoglobulin type E (IgE) antibodies. We have found very significant changes of the peptide profiles and a progressive reduction in residual-intact proteins after applying high pressure during tryptic proteolysis of β-Lg and BSA and chymotryptic proteolysis of β-Lg, α-La and BSA. A statistically significant decrease of the residual immunochemical reactivities of β-Lg tryptic and α-La chymotryptic hydrolysates prepared under high pressure, in comparison with the control samples hydrolysed at atmospheric pressure, was also observed.

Beran M., Urban M., Adamek L., Drahorad J., Molik P., Spevacek* J., Synytsya* A.
Simple and cheap isolation and purification of alkali-soluble β-glucans from Pleurotus spp. and Lentinus edodes mushrooms.
Proceedings of the 5th International Mushroom Conference. 5th – 8th September 2009. Nantong, China, 325-330.
Beside their traditional food usage, mushrooms can be used as a source of many pharmacologically active compounds, especially polysaccharides. Numerous bioactive polysaccharides or polysaccharide-protein complexes from medicinal mushrooms are described that appear to enhance innate and cell-mediated immune response and exhibit antitumor and other positive physiological activities in animals and humans. Bioactive mushroom polysaccharides differ greatly in their chemical composition and configuration. Most of the mushroom polysaccharides are 1,3;1,6-β-D-glucans with various contents of β-1,6 linkages. There is a broad similarity in the various methods that have been developed to extract the glucan polysaccharides from mushroom fruit-bodies, mycelium and liquid media. The extraction methods are rather complex and consist of several steps. The extracted fractions usually have to be further purified. We have developed a new, simple, cheap and ecological method of extraction of alkali-soluble β-glucans from Pleurotus spp. and Lentinula edodes mushrooms. Furthermore, this procedure consisting of two simple steps can be easily scaled-up to an industrial process. In the first step, mushroom biomass is hydrolyzed in a diluted aqueous solution of hydrochloric acid under heating. A significant part of mushroom biopolymers - proteins, acid-soluble polysaccharides, nucleic acids, and lipids - is hydrolyzed and solubilized under the treatment. The product containing the alkali-insoluble β-glucans was obtained in form of a segregated hydrogel upper layer of sediment after centrifugation of the acid-hydrolyzed mushroom suspension. Chitin and insoluble skeletal β-glucans can be separated from the remaining sediment in a subsequent step by alkali hydrolysis or another method described in literature. FTIR and 13C NMR analysis of the mushroom β-glucan preparations confirmed that the main polysaccharide component of all of them is branched β-(1,3)(1,6)-glucan. Pleurotus and shiitake glucans have quite similar structure. All products, however, contained detectable amounts of impurities, mainly proteins and lipids. Relative contents of these compounds in the preparations depend on the state of raw material, i.e. dried or fresh fruit bodies, and on the mushroom species.

Perlín.C., Svoboda K.
Analysis of sources of emitted substances in food processing industries.
Research report of project NVP II 2B08017.
The results of the analysis of emission sources and the values of these emissions from technological nodes are presented in a tabular form. Emission sources (except for boilers and sewage treatment facilities, which are not covered by this project) from six food processing industries (sugar refineries, poultry abattoirs and processing, slaughterhouses and meat processing factories, dairy industry, breweries and vegetable oil industry) were examined. Altogether 20 charts with emission sources and 24 charts with values of emitted substances are presented. In addition to collecting the basic values on emission production from food processing facilities in the Czech Republic, the performed analysis also proved differences in the approach of applicants and indulgence of administrative authorities; this caused the uncoordinated presentation of data, the use of different units, one’s own adaptations of applications against the valid legislation and also the inconsistent listing of emission sources within the same industrial branch.

Heroldova*M., Houska M., Vavrova*H., Kucera*P., Setinova*I., Honzova*S., Kminkova M., Strohalm J., Novotna P., Proskova A. (2009)
Influence of high pressure treatment on allergenicity of rDau c1 and carrot juice demonstrated by in vitro and in vivo tests.
High Pressure Research. 2009; 29(4): 695-704, ISSN 0895-7959 (print), 1477-2299 (online).
The aim of our study was to detect the influence of high pressure treatment (HPT) on allergenicity of recombinant allergen rDau c1 and carrot juice using in vitro and in vivo tests. The buffer solution of recombinant main carrot allergen r Dau c1 for basophil activation test (BAT) and Western blot (WB) was used. Dau c1 was pre-treated by pressure 500 MPa for 10 minutes and different temperatures (30, 40, 50°C) and pressure from 400 to 550 MPa for 3 and 10 minutes. Neither the HPT from 400 to 550 MPa for 3 and 10 minutes nor the HPT at 500 MPa for 10 minutes and temperature 30, 40 and 50°C had the influence on basophil activation by rDau c1. Serum samples of birch pollen allergic patients reacted in WB with solution of rDau c1 treated by HP 500 MPa for 10 minutes at temperature 30, 40 and 50°C. This pressure procedure did not influence the immune reactivity of rDau c1 in WB test. The structural changes of rDau c1 caused by HPT studied by circular dichroismus (CD) spectra were found. Mild increase of beta-helical structure was observed. The main changes were seen in rDau c1 samples treated 10 min. at 500 MPa and temperature 50°C. The influence of HPT on allergenicity of carrot juice was afterward studied. The reactivity in skin prick tests (SPT) and BAT did not show any influence of HPT on allergenicity of carrot juice. Pressure of 500 MPa for 10 minutes and temperature 30, 40 and 50°C did not inactivate allergen Dau c1 in carrot juice in WB. HPT from 450 to 550 MPa for 3 and 10 minutes at temperature 30°C had no influence on immune reactivity of Dau c1 in carrot juice. 19 patients underwent double-blind, placebo-controlled food challenge (DBPCFC). 13 of them reacted to placebo and were excluded, 1 patient did not react at any material (placebo, HPT material and fresh frozen carrot juice), 3 patients had positive test (reacted on HPT material and non-treated fresh frozen carrot juice) and 2 patients had negative reaction (reacted only on fresh frozen material). We did not confirm the influence of HPT on allergenicity of rDau c1 and carrot juice using in vitro and in vivo tests.

Cermak*P., Landfeld A., Mericka*P., Houska M.
Enterococcus faecium Growth Model.
Czech Journal of Food Sciences. 2009; 27(5): 361-371, ISSN 1212-1800.
Enterococci are bacteria commonly found in humans. However, the bacteria can cause severe infections in susceptible individuals. Strains of Enterococcus faecium have demonstrated increasing resistance to antibiotics, which is considered an important virulence factor. The contribution of E. faecium to infection-related illnesses has recently increased, and comprises most of the isolated Vancomycin-Resistant Enterococcus (VRE) strains. Enterococci are common contaminants of human milk processed in milk banks, and consumption of contaminated milk could cause severe infection-related complications if the checking mechanisms failed to detect such contamination. Extensive data are available on growth curves for E. faecium on growth in bouillon at pH values between 6 and 7, at temperatures of 5 to 20 °C, and for water activity values of a_w 0.97 to 0.997. These growth curves were replaced with non-linear Gompertz curves for microorganism growth, the parameters of which were correlated to temperature and pH values. A mathematical relationship to water activity could not be established as only two water activity levels had been used in experiment and the resulting model would be highly inaccurate. The issue of water activity was resolved by development of two separate models, one for each of the water activity values. The models correspond very well with the experimental growth curve data from which they had been developed. The model for water activity level 0.997 was used to predict the growth of E. faecium in cow and human milk (these two fluids have practically identical water activity), and the prediction was compared to experimental data. A good agreement was achieved between the prediction and experimental data in the case of cow milk. In the case of human milk, the model usually predicted a more rapid growth rate than was seen experimentally. The model was thus on the conservative side in all cases. Inhibitory agents naturally present in human milk might cause the slower growth rates.

Houska M., Heroldova*M., Vavrova*H., Kucera*P., Setinova*I., Havranova*M., Honzova*S., Strohalm J., Kminkova M., Proskova A., Novotna P. (2009)
Is the high-pressure treatment able to modify the allergenicity of main apple juice allergen Mal d1?
High Pressure Research 2009; 29(1): 14-22, ISSN 0895-7959 (print), 1477-2299 (online).
The aim of this paper is to study the influence of high-pressure treatment (HPT) on the structure and allergenicity of the recombinant main apple allergen rMal d1, apple juice and homogenates prepared from the Golden delicious variety. In vitro and in vivo tests have been used for evaluation of HPT effects: circular dichroism (CD) spectroscopy, the basophile activation test (BAT – CD63 expression) and the Western blot (WB) with IgE positive serum, skin prick-to-prick test (PTP). Apple homogenates were tested using a double blind placebo controlled food challenge (DBPCFC). rMal d1 solutions treated at 500 MPa for 10 minutes at 30°C showed the greatest changes in CD spectra, when compared to untreated samples. The WB test of these solutions showed that HPT did not influence allergic reactions. The BAT also failed to differentiate between HP treated and untreated rMal d1 solutions. Apple juice treated with HPT in the range of 450-550 MPa for 3 and 10 minutes at 30°C failed to show any differences in BAT and PTP tests compared with untreated juice. WB tests of apple juice showed that bands of proteins corresponding to the Mal d1 standard are not affected by HPT. In DBPCFC with apple homogenates, only 14 of 20 patients could be evaluated; 5 patients failed to react to the HPT homogenate but did react to the untreated homogenate (35.7%), 5 patients reacted to both the HPT homogenate and the untreated homogenate (35.7%) and 4 (28.6%) failed to react to either. HPT using the stated parameters was not capable of altering the allergenicity of rMal d1 in pure solutions in our group of patients. Also, allergenicity of apple juice and apple homogenates, prepared using this technique, cannot be substantially decreased.

A results and technology with practical applications.

A simple enzymatic test to determine milk allergen in foods.
Validated technology (Sedium, s.r.o.).
The test is intended for consumers dependent on milk free diet, producers, distributors and vendors of milk free foodstuffs. It does not require any special equipment and makes it possible to confirm or exclude the presence of milk in a foodstuff. Determination is based on the reaction of the most important milk allergen, beta-lactoglobulin, with a specific enzyme-marked antibody. The test concludes itself with a change in colour, the intensity of the colour being proportional to the quantity of allergen contained in the sample. The test makes it possible to examine five samples simultaneously within one hour, and can be easily performed and evaluated both in home and industrial environments. The kit contains all the items needed for successfully conducting the test. Negative and positive reference samples are enclosed and are tested concurrently with the samples of analyzed foods.

A simple enzymatic test to determine gluten in foods.
Validated technology (Sedium, s.r.o.).
The test is intended for consumers coping with celiac disease, for producers, distributors and vendors of gluten free foods. It does not require any special equipment and makes it possible to confirm or exclude the presence of gluten in a foodstuff. Determination is based on the reaction of gluten with specific antibodies. The test concludes itself with a change in colour, the intensity of the colour being proportional to the quantity of gluten contained in the sample. The test makes it possible to examine five samples simultaneously within one hour.

A validated preparation technology of fresh carrot juice without active allergen Dau c1.
Validated technology (Beskyd Fryčovice s.r.o.).
Fresh carrot juice without the active allergen Dau c1, distinguishing itself by the content of carrot matter with the admixture of fresh apple juice with the authentic content of natural catalysts of oxidative processes.

A validated production technology of deallergized apple juice.
Validated technology (Beskyd Fryčovice s.r.o.).
A validated production technology of deallergized apple juice, including the verification of its shelf life from the viewpoint of its microbial and sensory quality, the deallergization effect and the stability of vitamin C, using cold pasteurization by high pressure for microbial stabilization.

Protein mix for food purposes and the way of its production.
Validated technology (spol.Adler Wellness Produkte s.r.o., ČR).
The protein mix for food purposes, based on egg white containing solid fibre, is composed of dried egg white and a mix of saccharides, solid fibre, hydrocolloids, food additives and water. Its production consists in homogenizing the mix of weighed dry ingredients, adding water and another homogenization. After some standing time the mix is kneaded and then pieces of the required shape and size are formed with help of a pressing appliance and a mould. The final shape is stabilized using coagulation in a hot water bath.

Production of food supplement “Imunocomplex”.
The production of raw material based on cell walls of yeast biomass for the manufacture of the immunostimulative food supplement “Imunocomplex” by Nexar Co.

Patents and utility models listed in the overall survey of results are ready for practical applications – see section Publication.

Patents and Utility models

Patent č. 300164 (2009): Preservation method of fresh hop. (Houška M., Strohalm J., Krofta*K., Mikyška*A.)

Patent č. 300200 (2009): Method of separating protein fraction (Jelínek*M., Kolomazník*K., Jelínek*J., Beran M., Adámek L., Urban M.)

Patent č. 300201(2009): Method of making food supplement (Jelínek*M., Beran M., Adámek L., Urban M.)

Patent č. 300210 (2009): Method of removing sugar from egg white and gumbo. (Adámek L., Kýhos K., Beran M., Rutová E.)

Patent č. 300697( 2009): Functional symbiotic edible food packages. (Urban M., Beran M., Adámek L., Molík P.)

Užitný vzor reg. č. 19230 (2009): Rutin-containing milling grain mixture. (Paulíčková I.)

Užitný vzor reg. č. 19427 (2009): Immuno-analytic kit for determination of mustard proteins in foodstuffs. (Gabrovská D., Rysová J., Hanák P., Nesládková K., Hušková M., Štumr*F., Tomková*K., Šafářová*P., Dvorská*P.)

Užitný vzor reg. č. 19483 (2009): Combined protein for food processing. (Kýhos K.)

Užitný vzor reg. č. 19556 (2009): Hypoallergenic bacterial culture for preparing fermented functional foods. (Pechačová*M., Němečková*I., Roubal*P, Gabrovská D.)

Užitný vzor reg. č. 19629 (2009): Bakery product containing rutin. (Paulíčková I.)

Užitný vzor reg. č. 19640 (2009): Device for processing combined protein under water surface. (Kýhos K., Strohalm J.)

Užitný vzor reg. č. 19674 (2009): Butter or dressing with addition of vegetable raw materials. (Němečková*I., Pechačová*M., Roubal*P., Gabrovská D.)

Užitný vzor reg. č. 19675 (2009): Cottage cheese dessert with vegetable flour. (Němečková*I., Pechačová*M., Roubal*P., Gabrovská D.)

Užitný vzor reg. č. 19676 (2009): Jelly butter with psyllium. (Němečková*I., Pechačová*M., Roubal*P., Gabrovská D.)

Užitný vzor reg. č. 19703 (2009): Buckwheat-barley beer. (Prokeš*J., Škach*J., Gabrovská D., Ouhrabková J., Paulíčková I.)

Užitný vzor reg. č.19733 (2009): Food supplement. (Hromádka*R., Beran M., Zadák*Z.)

Užitný vzor reg. č. 19855 (2009): Food with Tartarian buckwheat. (Rysová J., Paulíčková I., Janovská*D., Ouhrabková J., Gabrovská D.)

Užitný vzor reg. č. 19868 (2009): Mixture for food with Tartarian buckwheat. (Rysová J., Paulíčková I., Janovská*D., Ouhrabková J., Gabrovská D.)

Užitný vzor reg.č. 20005 (2009).: Composition for stimulation growth of cells in tissue cultures. (Mejsnar*J., Houška M., Čejková*P.)

Užitný vzor reg.č. 20021 (2009): Gelatin. (Kučera J., Prošková A.)

Užitný vzor reg.č. 20022 (2009): Gelatin. (Kučera J., Prošková A., Škarková*L.)

Prototype, Functional sample

A pilot apparatus for food decontamination using ozone water. (Landfeld A., Strohalm J., Kýhos K.)