Person: Martín De Santos, María Del Rosario
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First Name
María Del Rosario
Last Name
Martín De Santos
Affiliation
Universidad Complutense de Madrid
Faculty / Institute
Veterinaria
Department
Nutrición y Ciencia de los Alimentos
Area
Nutrición y Bromatología
Identifiers
14 results
Search Results
Now showing 1 - 10 of 14
Item PCR-based assay for the detection of Alternaria species and correlation with HPLC determination of altenuene, alternariol and alternariol monomethyl ether production in tomato products(Food Control, 2012) Pavón, Miguel Ángel; Luna, Agustín; Cruz, Silvia de la; González Alonso, María Isabel; Martín De Santos, María Del Rosario; García Lacarra, TeresaAlternaria spp. contamination and subsequent production of mycotoxins is a common problem in vegetable crops. Identification of Alternaria species by traditional methods requires specific skills and may not detect toxigenic moulds inactivated by food processing. By using molecular methods such as PCR the detection of Alternaria spp. becomes possible directly from the food or feed samples. In this study, a PCR method based on the Internal Transcribed Spacer (ITS) genetic marker has been used for detection of Alternaria spp. in raw and processed commercial tomato samples. Occurrence of altenuene, alternariol and alternariol methyl ether in the samples was analysed by high-performance liquid chromatography (HPLC) in order to assess the ability of the PCR assay to identify tomato samples containing Alternaria mycotoxins. The PCR assay revealed the presence of Alternaria spp. DNA in 41 out of 90 commercial samples (45.6%), while HPLC detected at least one of the Alternaria mycotoxins within 31 of the PCR positive samples. Detection of Alternaria DNA correlated well with the presence of the analysed Alternaria mycotoxins, indicating that the PCR protocol developed in this work for detection of Alternaria spp. DNA could be used as an indirect marker of the presence of Alternaria mycotoxins in raw and processed tomato products.Item A sensitive and specific real-time PCR targeting DNA from wheat, barley and rye to track gluten contamination in marketed foods(LWT- Food Science and Technology, 2019) Sohrabi, v; Cruz, Silvia de la; García García, Aina; Madrid García, Raquel; García Lacarra, Teresa; Martín De Santos, María Del Rosario; González Alonso, María IsabelThis work reports a real-time PCR assay to specifically detect the presence of DNA from the main gluten-containing cereals wheat, barley and rye (WBR) in foodstuffs. The WBR real-time PCR uses two specific primers and a Taqman probe for amplification of a 118 bp DNA fragment common to the three target cereals on the ribosomal internal transcribed spacer (ITS) region. In-house validation of the method was performed employing three binary arrays containing different concentrations (100 000–10 mg/kg) of a wheat/barley/rye mixture in maize flour: untreated, soft heat-treated at 160 °C/13 min and intense heat-treated at 200 °C/20 min. Results showed optimal PCR amplification efficiency, reproducibility and sensitivity with an overall limit of detection of 10–50 mg/kg of the target cereals, theoretically equating to approximately 1–5 mg/kg of gluten. Applicability of the assay was further assessed through a market analysis of 220 food products by the real-time PCR assay and the official R5-based ELISA. Comparative results highlighted the ability of the proposed methodology as a highly sensitive and robust procedure to detect the presence of potentially harmful concentrations of undeclared gluten-containing cereals in some of the tested samples, supporting results of the immunological assays currently used for gluten determination in foods.Item Duplex real-time PCR method for the detection of sesame (Sesamum indicum) and flaxseed (Linum usitatissimum) DNA in processed food products(Food Additives and Contaminants - Part A Chemistry, Analysis, Control, Exposure and Risk Assessment, 2015) López-Calleja, Inés María; Cruz, Silvia de la; Martín De Santos, María Del Rosario; González Alonso, María Isabel; García Lacarra, TeresaThe development of a duplex real-time polymerase chain reaction (PCR) method allowing the simultaneous detection of sesame and flaxseed DNA in commercial food products is described. This duplex real-time PCR technique is based in the design of sesame- and flaxseed-specific primers based on the ITS1 region and two TaqMan fluorescent probes. The method was positive for sesame and flaxseed, and showed no cross-reactivity for all other heterologous plant and animal species tested. Sesame and flaxseed could be detected in a series of model samples with defined raw and heat-treated sesame in flaxseed, and flaxseed in sesame, respectively, with detection limits of 1.3 mg kg(-1) for sesame and 1.4 mg kg(-1) for flaxseed. The applicability of the assay for determining sesame and flaxseed in different food matrices was investigated by analysing a total of 238 commercial foodstuffs. This PCR method is useful for highly selective and sensitive detection of traces of sesame and flaxseed in commercial food products.Item Survey of undeclared allergenic pistachio (Pistacia vera) in commercial foods by hydrolysis probe real-time PCR(Food Control, 2014) López-Calleja, Inés María; Cruz, Silvia de la; González Alonso, María Isabel; García Lacarra, Teresa; Martín De Santos, María Del RosarioTree nut allergies represent an important health problem in industrialized countries. Among these, pistachio (Pistacia vera) kernels which are consumed as snack foods and used as ingredients in confectionery, chocolates, meat products, and ice-cream industries have been reported to cause IgE-mediated allergic reactions. Trace amounts of undeclared pistachio allergens can cause serious health risks for food-allergic consumers. In order to provide an appropriate method for the detection of pistachio in food products, a real-time polymerase chain reaction (PCR) system for the specific and sensitive detection of pistachio was developed. The sensitivity was investigated on spiked wheat flour samples with defined raw and heat-treated pistachio contents (0.1–100,000 mg/kg). The real-time PCR detected pistachio in these mixtures down to the lowest investigated spike level of 0.1 mg/kg. In addition, analysis of different retail samples from the market was performed to demonstrate the suitability of the assay in the food industry. The real-time PCR results obtained from the analysis of 229 commercial food products revealed 29 that didn't declare pistachio or traces on the label but were found to contain pistachio. The presented real time PCR method is useful for relatively fast, highly selective, and sensitive detection of pistachio in food samples.Item Identification and characterisation of the proteins bound by specific phage‐displayed recombinant antibodies (scFv) obtained against Brazil nut and almond extracts(Journal of the Science of Food and Agriculture, 2017) Cruz. Silvia de la; Madrid García, Raquel; García García, Aina; Alcocer, Marcos; Martín De Santos, María Del Rosario; González Alonso, María Isabel; García Lacarra, TeresaBACKGROUND: Almonds and Brazil nuts are widely consumed allergenic nuts whose presence must be declared according to food labelling regulations. Their detection in food products has been recently achieved by ELISA methods with recombinant antibodies (scFv) isolated against complete Brazil nut and almond protein extracts. The screening of phage-scFv libraries against complete protein extracts confers a series of advantages over the use of purified proteins, as recombinant proteins might alter their native folding. However, using this strategy, the nature of the target detected by phage-displayed antibodies remains unknown, and requires further research to identify whether they are nut allergens or other molecules present in the extract, but not related to their allergenic potential. RESULTS: Electrophoretic, chromatographic, immunological and spectrometric techniques revealed that the Brazil nut (BE95) and almond (PD1F6 and PD2C9) specific phage-scFvs detected conformational epitopes of the Brazil nut and almond 11S globulins, recognised by WHO/IUIS as Ber e 2 and Pru du 6 major allergens. Circular dichroism data indicated that severe heat treatment would entail loss of epitope structure, disabling scFv for target detection. CONCLUSIONS: The presence of important Brazil nut and almond allergens (Ber e 2 and Pru du 6) in foodstuffs can be determined by using phage-display antibodies BE95, PD1F6 and PD2C9 as affinity probes in ELISA.Item The use of high‐performance liquid chromatography to detect ochratoxin A in dried figs from the Spanish market(Journal of the Science of Food and Agriculture, 2011) Pavón, Miguel Ángel; González Alonso, María Isabel; Cruz, Silvia de la; Martín De Santos, María Del Rosario; García Lacarra, TeresaBACKGROUND: Detection and quantification of ochratoxin A (OTA) in dried fig samples purchased in Spain has been carried out using high-performance liquid chromatography with fluorescence detection after extraction with methanol and sodium bicarbonate, and clean-up by using an immunoaffinity column. RESULTS: The detection limit of the method was 0.06 ng/g, and the limit of quantification 0.18 ng/g. OTA was detected in 31 (88.6%) out of 35 samples of dried figs analysed, with concentrations that ranged from < 0.1 to 277 ng/g. However, only three samples contained OTA concentrations above the tolerable level set by European Commission regulations for dried vine fruits (10 ng/g). CONCLUSION: The results of this survey show the value of monitoring OTA in dried figs especially if they are home grown.Item Development of real-time PCR assays to detect cashew (Anacardium occidentale) and macadamia (Macadamia intergrifolia) residues in market analysis of processed food products(LWT-Food Science and Technology, 2015) López-Calleja, Inés María; Cruz, Silvia de la; González Alonso, María Isabel; García Lacarra, Teresa; Martín De Santos, María Del RosarioReal-time polymerase chain reaction (PCR)-based assays for detection of cashew (Anacardium occidentale) and macadamia nut (Macadamia intergrifolia) traces in food products are described here. The real time PCR technique proposed herein were developed based on the design of macadamia and cashew-specific primers from the ITS region and a TaqMan fluorescent probe. The methods were positive for cashew and macadamia respectively, and negative for all other heterologous plant and animal species tested. Using a series of model samples with defined levels of raw and heat-treated cashew and macadamia nut, respectively, within a range of concentrations of 0.1–100 000 mg kg−1, practical detection limits of 0.1 mg kg−1 for cashew and macadamia nut were estimated. Practical applicability of the PCR methods was tested by the analysis of a total of 214 commercial foodstuffs. These PCR methods, are useful for highly selective, and sensitive detection of traces of macadamia and cashew nuts in commercial food products and is therefore proposed as a ready-to-use analytical tool to trace these target allergens in foods.Item Development of a real time PCR assay for detection of allergenic trace amounts of peanut (Arachis hypogaea) in processed foods(Food Control, 2013) López-Calleja, Inés María; Cruz, Silvia de la; Pegels, Nicolette; González Alonso, María Isabel; García Lacarra, Teresa; Martín De Santos, María Del RosarioPeanut allergic reactions can result from the ingestion of even very small quantities of peanut and represent a severe threat to the health of sensitized individuals. In order to protect the allergic consumer, efficient and reliable methods are required for the detection of allergenic ingredients. For this purpose, we have developed a TaqMan real-time PCR method for specific detection of peanut in commercial food products. The assay uses two genetic makers in the Arah2 (125 bp) and ITS1 (90 bp) gene respectively, and TaqMan probes. The nuclear 18S rRNA gene was employed as a positive amplification control. Results obtained on sensitivity of both Arah2 and ITS primers with mixtures spiked with different concentrations of the target showed detection levels of 10 and 0.1 ppm, respectively. The applicability of the real-time PCR protocol to detect the presence of peanut DNA in commercial food products was determined through analysis of 123 different commercial products with the ITS primers which aimed higher sensitivity than Arah2 primer pair. The performance of the real-time PCR proposed herein allows a highly sensitive detection of peanut DNA in all the different food products, which declared to contain peanut material as well as in others were the presence of peanut or its traces wasn't declared in the labeling.Item Duplex real-time PCR using TaqMan® for the detection of sunflower (Helianthus annuus) and poppy (Papaver rhoeas) in commercial food products(LWT - Food Science and Technology, 2015) López-Calleja, Inés María; Cruz, Silvia de la; González Alonso, María Isabel; García Lacarra, Teresa; Martín De Santos, María Del RosarioThe paper presents a duplex real-time PCR assay for the simultaneous detection of traces of potentially allergenic sunflower and poppy seeds in commercial food products. For the design of sunflower and poppy-specific primers and two TaqMan fluorescent probes labeled with FAM and HEX respectively, the ITS1 region was selected. The duplex real-time PCR assay did not show any cross-reactivity for all other heterologous plant and animal species tested commonly used in the food industry. Analysis of serially diluted raw and heat-treated sunflower in poppy with a range of concentrations of 0.1–100,000 mg kg−1 and raw and heat-treated poppy in sunflower with the same concentrations resulted in good linearity, with a LOD of 0.1 mg kg−1 of sunflower and poppy content. The duplex real-time PCR assay was applied to verify correct labeling of 238 commercial foodstuffs comprising: nut bars, cookies, chocolates, creams, bonbons, cereals, bread, ice cream, meat products and prepared food products. Results indicate that this duplex PCR is highly sensitive and selective, which makes it suitable for the detection of sunflower and poppy traces in food samples. In addition, it can be useful for monitoring the effectiveness of cleaning processes in the production units of the food industry.Item Isolation of recombinant antibody fragments (scFv) by phage display technology for detection of almond allergens in food products(Food Control, 2015) Cruz, Silvia de la; Cubillos-Zapata, Carolina; López-Calleja, Inés María; Ghosh, Satyabrata; Alcocer, Marcos; González Alonso, María Isabel; Martín De Santos, María Del Rosario; García Lacarra, TeresaTree nut allergies are considered an important health issue in developed countries. To comply with the regulations on food labeling, reliable allergen detection methods are required. In this work we isolated almond-specific recombinant antibody fragments (scFv) from a commercial phage display library bypassing the use of live animals, hence being consistent with the latest policies on animal welfare. To this end an iterative selection procedure employing the Tomlinson I phage display library and a crude almond protein extract was carried out. Two different almond-specific scFv (named PD1F6 and PD2C9) were isolated after two rounds of biopanning, and an indirect phage ELISA was implemented to detect the presence of almond protein in foodstuffs. The isolated scFvs demonstrated to be highly specific and allowed detection of 40 ng mL−1 and 100 ng mL−1 of raw and roasted almond protein, respectively. The practical detection limit of the assay in almond spiked food products was 0.1 mg g−1 (110–120 ppm). The developed indirect phage ELISA was validated by analysis of 92 commercial food products, showing good correlation with the results obtained by a previously developed real-time PCR method for the detection of almond in foodstuffs. The selected phage clones can be affinity maturated to improve their sensitivity and genetically engineered to be employed in different assay formats.