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Öğe Amanitin and phallotoxin concentration in Amanita phalloides var. alba mushroom(Pergamon-Elsevier Science Ltd, 2013) Kaya, Ertuğrul; Yılmaz, İsmail; Sinirlioğlu, Zeynep Aydın; Karahan, Selim; Bayram, Recep; Yaykaşlı, Kürşat Oğuz; Severoğlu, ZekiAlthough rarely seen, Amanita phalloides var. alba, a variety of A. phalloides type mushrooms, causes mushroom poisoning resulting in death. Since it is frequently confused with some edible mushrooms due to its white colored cap and macroscopic appearance, it becomes important in toxicological terms. Knowledge of the toxin amount contained in this mushroom type is invaluable in the treatment of cases involving poisoning. In this study, we examined the toxin levels of various parts of the A. phalloides var. alba mushroom growing Duzce region of Turkey. Toxin analyses were carried out for A. phalloides var. alba, which were collected from the forests Duzce region of Turkey in 2011, as a whole and also separately in its spore, pileus, gills, stipe and volva parts. The alpha amanitin, beta amanitin, gamma amanitin, phalloidin and phallacidine analyses of the mushrooms were carried out using the RP-HPLC method. A genetic analysis of the mushroom showed that it had similar genetic characteristics as A. phalloides and was a variety of it. The lowest toxins quantity was detected in spores, volva and stipe among all parts of the mushroom. The maximum amount of amatoxins was measured in the gills. The pileus also contained a high amount of amatoxins. Generally, amatoxins and phallotoxin concentrations were lower as compared to A. phalloides, but interestingly all toxins other than gamma toxin were higher in the spores of A. phalloides var. alba. The amount of toxin in all of its parts had sufficient concentrations to cause death. With this study, the amatoxin and phallotoxin concentrations in A. phalloides var. alba mushroom and in its parts have been revealed in detail for the first time. (C) 2013 Elsevier Ltd. All rights reserved.Öğe Amatoxin and phallotoxin concentration in Amanita phalloides spores and tissues(Sage Publications Inc, 2015) Kaya, Ertuğrul; Karahan, Selim; Bayram, Recep; Yaykaşlı, Kürşat Oğuz; Çolakoğlu, Serdar; Sarıtaş, AyhanMost of the fatal cases of mushroom poisoning are caused by Amanita phalloides. The amount of toxin in mushroom varies according to climate and environmental conditions. The aim of this study is to measure -, -, and -amanitin with phalloidin and phallacidin toxin concentrations. Six pieces of A. phalloides mushrooms were gathered from a wooded area of Duzce, Turkey, on November 23, 2011. The mushrooms were broken into pieces as spores, mycelium, pileus, gills, stipe, and volva. -, -, and -Amanitin with phalloidin and phallacidin were analyzed using reversed-phase high-performance liquid chromatography. As a mobile phase, 50 mM ammonium acetate + acetonitrile (90 + 10, v/v) was used with a flow rate of 1 mL/min. C18 reverse phase column (150 x 4.6 mm; 5 mu m particle) was used. The least amount of -amanitin toxins was found at the mycelium. The other toxins found to be in the least amount turned out to be the ones at the spores. The maximum amounts of amatoxins and phallotoxin were found at gills and pileus, respectively. In this study, the amount of toxin in the spores of A. phalloides was published for the first time, and this study is pioneering to deal with the amount of toxin in mushrooms grown in Turkey.Öğe Clinical importance of toxin concentration in Amanita verna mushroom(Pergamon-Elsevier Science Ltd, 2014) Yılmaz, İsmail; Kaya, Ertuğrul; Sinirlioğlu, Zeynep Aydın; Bayram, Recep; Sürmen, Mustafa Gani; Çolakoğlu, SerdarPoisoning from Amanita group of mushrooms comprises approximately 3% of all poisonings in our country and their being responsible for nearly the entire fatal mushroom poisonings makes them important. These mushrooms contain primarily two types of toxins, amatoxins and phallotoxins. Phallotoxins have a more limited toxicity potential and they primarily consist of phalloidin (PHN) and phallacidin (PCN). Amatoxins, on the other hand, are very toxic and they primarily consist of alpha-amanitin (AA), beta-amanitin (BA) and gamma-amanitin (GM. Toxin levels can vary among various species, even among varieties of the same species, of Amanita mushroom family. Revealing the differences between the toxin compositions of the Amanita species that grow in our region may contribute to the clinics of poisonings. Our study aims at showing in detail the toxin levels in various parts of Amanita verna mushroom. A. verna mushrooms needed for toxin analysis were collected from Kozak Plateau near Ayvalik county of Balikesir, Turkey in April 2013. The mushrooms were divided into their parts as pileus, gills, stripe and volva. Following the procedures required before the analysis, the AA, BA, GA, PHN and PCN levels were measured using the RP-HPLC method. While the lowest level of amatoxin was in the volva of the mushroom, the highest was measured in the gills. This was followed by pileus and stripe where the levels were close to each other. Similarly, the highest level of phallotoxin was measured in the gills. Gamma toxin and phalloidin were at lower amounts than the other toxins. A. verna is frequently confused with edible mushrooms with white caps due to its macroscopic similarity. If just one of them is eaten by mistake by an adult person with no mushroom experience, it can easily poison them. The amount of amatoxin is more as compared to Amanita phalloides and A. phalloides var. alba. Particularly, the AA and BA levels are approximately three times higher, whereas GA levels are lower. Similarly, the level of PCN is approximately four times higher as compared to A. phalloides and A. phalloides var. alba; by contrast, the level of PNH is about a half of theirs. In summary, it can be said that A. verna is a more toxic mushroom than A. phalloides and has a higher rate of mortality. With our study, the amatoxin and phallotoxin concentrations and distribution in A. verna mushrooms were shown in detail for the first time and it would be useful to carry out more similar studies with other members of Amanita family growing in various parts of the world. (C) 2014 Elsevier Ltd. All rights reserved.Öğe Düzce Yöresinde Yetişen Amanita Phalloides Mantarındaki Alfa Amanitin Düzeyinin Hplc Yöntemiyle Ölçümü(2012) Kaya, Ertuğrul; Karahan, Selim; Hancı, Mustafa; Yaykaşlı, Kürşat Oğuz; Sarıtaş, Ayhan; Bayram, Recep; Arslan, Seyfullah OktayAmaç: Düzce ili sınırlarında 2010 yılında toplanan Amanita phalloides mantarındaki alfa amanitin toksin düzeyinin HPLC yöntemiyle ölçümü amaçlanmıştır. Yöntem: Bir mantar bütün olarak, diğeri ise parçalara ayrılarak ekstraksiyon yapılmıştır. Ölçümler HPLC cihazında 303 nm UV dalga boyu, 250x4,6 mm C18 5 µm partikül içeren kolon kullanılarak gerçekleştirilmiştir. Mobil faz olarak amonyum asetat metanol asetonitril (801010, v/v/v) kullanılmış ve akış hızı 1 mL/dakikaya ayarlanmıştır. Sonuçlar 1 g kuru mantardaki toksin miktarı olarak verilmiştir. Bulgular: Bütün mantardaki alfa amanitin miktarı 4,806 mg (0,033), şapkada 3,522 mg (0,024), lamelde 5,318 mg (0,056), halkada 0,903 mg (0,004), sapta 2,577 mg (0,037), kapçıkta 0,698 mg (0,008) olarak ölçüldü. Sonuç: Düzce yöresinde yetişen Amanita phalloides mantarlarındaki alfa amanitin düzeyleri, başka bölgelerde yetişenlerden farklılık göstermektedir. Bulduğumuz sonuçlardan daha yüksek ve daha düşük seviyede toksin düzeyi ölçülmüş araştırmalar literatürde mevcuttur. Bu farklılığın etkenleri arasında iklim şartları yanında ekstraksiyon ve analiz yöntemlerindeki farklılıklar da rol oynayabilir.Öğe Effects of erdosteine on alpha amanitin-induced hepatotoxicity in mice(Taylor and Francis Ltd, 2016) Kaya, Ertuğrul; Yılmaz, İsmail; Admış, Özlem; Oktay, Murat; Bayram, Recep; Bakırcı, Sinan; Çolakoğlu, SerdarThe aim of this study was to investigate beneficial effects of erdosteine in the alpha amanitine-induced hepatotoxicity in mice. Three hours after giving alpha amanitin (0.5 mg/kg, i.p.) to the mice, they were administered silibinin (50 mg/kg/d, i.p.) or erdosteine (100 mg/kg/d, oral) therapies once a day for 3 d. A histopathological examination of their liver tissues was carried out 24 h after the last treatment; transaminase levels, blood urea nitrogen, urea, and creatinine were analyzed in serum. Erdosteine showed a beneficial effect by significantly improving the functional parameters particularly in alpha amanitin-induced hepatotoxicity and partially in renal toxicity. In the histopathological evaluation, the toxicity that was generated with alpha amanitin was significantly reduced by erdosteine, showing a possible hepatoprotective effect. © 2016 Informa UK Limited, trading as Taylor & Francis Group.Öğe Erdosteine reduces cytotoxicity induced by alpha- and beta-amanitin, but not gamma-amanitin, in CA3 hepatocyte cultures(Pergamon-Elsevier Science Ltd, 2022) Bayram, Recep; Yılmaz, İsmail; Yaykasli, Kursat Oguz; Kaya, ErtuğrulAmanitin poisoning still has no particular, effective antidote. Erdosteine has been shown to protect numerous tissues, particularly those in the liver. This study investigates the potential therapeutic effects of erdosteine on alpha-, beta-and gamma-amanitin-induced hepatotoxicity in in vitro models. Three hours after administering amatoxins at various concentrations (1-50 mu g/mL) to the cells of the C3A human hepatocyte cell line, erdosteine was administered in different concentrations (i.e., 1, 10, 50, 100 and 250 mu g/mL). The 3-(4,5-dimethylthiazol-2yl)-2,5-diphenyltetrazolium bromide (MTT) assay was selected to determine cell viability. When concentrations of 1, 10, 50, 100 and 250 mu g/mL of erdosteine were applied to cell lines, the following cell viability rates were obtained: 106%,99%,93%,86% and 86%, respectively, at a 10 mu g/mL alpha-amanitin-induced toxicity; 43%,41%,41%,37% and 35%, respectively, at a 25 mu g/mL alpha-amanitin-induced toxicity; 44%,42%,41%,39% and 41%, respectively, at a 50 mu g/mL alpha-amanitin-induced toxicity; 136%,142%,143%,137% and 120%, respectively, at a 10 mu g/mL beta-amanitin-induced toxicity; 113%,107%,107%,106% and 86%, respectively, at a 25 mu g/mL beta-amanitin-induced toxicity; 78%,77%,77%,74% and 70%, respectively, at a 10 mu g/mL gammaamanitin-induced toxicity; and 39%,40%,39%,35% and 31%, respectively, at a 25 mu g/mL gamma-amanitininduced toxicity. This study was the first to evaluate the in vitro efficacy of erdosteine in cytotoxicity induced by alpha-, beta-and gamma-amanitin. Non-high (low and medium) doses of erdosteine are capable of nearly entirely preventing toxicity at mild hepatotoxic concentrations caused by amatoxin and partially preventing toxicity at moderate and severe concentrations. The beneficial effects of erdosteine, especially on the toxicity of alpha-and beta-amanitin, are promising.Öğe Evaluation and comparison of alpha- and beta-amanitin toxicity on MCF-7 cell line(2014) Kaya, Ertuğrul; Bayram, Recep; Yaykaşlı, Kürşat Oğuz; Yılmaz, İsmail; Bayram, Sait; Yaykaşlı, Emine; Gepdiremen, Ali AkçahanBackground/aim: Alpha- and beta-amanitins are the main toxins of the poisonous Amanita phalloides mushroom. Although there are many studies available concerning alpha-amanitin, there are limited data about beta-amanitin in the literature. Therefore, this study is aimed at comparing the toxic effects of alpha- and beta-amanitin on the MCF-7 cell line. Materials and methods: The alpha- and beta-amanitins used for this research were purified from Amanita phalloides by preparative high-performance liquid chromatography. The MCF-7 breast cancer cell line was used, and specific concentrations of the toxins (100, 10, 1, 0.1, and 0.01 µg/mL) were applied to the cells. The MTT test was performed to determine the level of toxicity, and the quantity of protein in the cell was measured using the biuret test. Results: The alpha-amanitin showed a higher toxicity at 36 h, while the highest inhibition of protein synthesis by the beta-amanitin was observed at 24 h. Conclusion: It was shown that the beta-amanitin may be responsible for toxicity, like alpha-amanitin, in Amanita phalloides mushroom poisoning. The early inhibition of protein synthesis for beta-amanitin might be useful for future experiments and research.Öğe Evaluation and comparison of alpha- and beta-amanitin toxicity on MCF-7 cell line(Tubitak Scientific & Technical Research Council Turkey, 2014) Kaya, Ertuğrul; Bayram, Recep; Yaykaşlı, Kürşat Oğuz; Yılmaz, İsmail; Bayram, Sait; Yaykaşlı, Emine; Gepdiremen, Ali AkçahanBackground/aim: Alpha- and beta-amanitins are the main toxins of the poisonous Amanita phalloides mushroom. Although there are many studies available concerning alpha-amanitin, there are limited data about beta-amanitin in the literature. Therefore, this study is aimed at comparing the toxic effects of alpha- and beta-amanitin on the MCF-7 cell line. Materials and methods: The alpha- and beta-amanitins used for this research were purified from Amanita phalloides by preparative high-performance liquid chromatography The MCF-7 breast cancer cell line was used, and specific concentrations of the toxins (100, 10, 1, 0.1, and 0.01 mu g/mL) were applied to the cells. The MTT test was performed to determine the level of toxicity, and the quantity of protein in the cell was measured using the biuret test. Results: The alpha-amanitin showed a higher toxicity at 36 h, while the highest inhibition of protein synthesis by the beta-amanitin was observed at 24 h. Conclusion: It was shown that the beta-amanitin may be responsible for toxicity, like alpha-amanitin, in Amanita phalloides mushroom poisoning. The early inhibition of protein synthesis for beta-amanitin might be useful for future experiments and research.Öğe The evaluation and comparison of the alpha and beta amanitin toxicity on MCF-7 cell line(Current Biology Ltd, 2013) Kaya, Ertuğrul; Bayram, Recep; Yaykaşlı, Kürşat Oğuz; Yılmaz, İsmail; Bayram, Sait; Yaykaşlı, Emine; Gepdiremen, Ali Akçahan…Öğe Extracorporeal Shockwave Increases the Effectiveness of Systemic Antibiotic Treatment in Implant-Related Chronic Osteomyelitis: Experimental Study in a Rat Model(Wiley, 2014) İnanmaz, Mustafa Erkan; Uslu, Mustafa; Işık, Cengiz; Kaya, Ertuğrul; Taş, Tekin; Bayram, RecepImplant-related chronic osteomyelitis is a serious complication of orthopedic surgery requiring implant removal and radical debridement. Extracorporeal shockwave (ESW) have demonstrated significant bactericidal effectiveness in vitro and effectiveness and safety were evaluated in an animal model of osteomyelitis. In this experimental study, we aimed to test our hypothesis that the use of ESW together with systemic antibiotic treatment will provide synergy for the treatment of implant-related chronic osteomyelitis caused by methicillin-susceptible Staphylococcus aureus (MSSA). The proximal tibia of 32 rats was contaminated with (10)8CFU/ml methicillin-sensitive S. aureus (MSSA-ATCC 29213) and Kirschner-wires were placed into the medulla of the tibia. After 4 weeks, Kirschner-wires were removed and the rats were randomly divided into four groups: group I, untreated contaminated control group; group II, receiving only ESW therapy; group III, receiving only systemic teicoplanin; group IV, treated with a combination of ESW and systemic teicoplanin. ESW was applied twice to the infected limbs and all rats were sacrificed at the end of 8th week. The degree of tibial osteomyelitis was assessed by quantitative culture analysis. Bacterial counts in groups III and IV were significantly reduced relative to the control (p=0.002 and 0.001, respectively). The decrease in bacterial counts was more pronounced and significant in group IV compared to group III (p=0.024). In group II, bacterial counts also decreased, but the differences were in significant (p=0.068). Our experimental model suggests that ESW provides significant synergy for systemic antibiotic treatment. However, further clinical trials are required in order to use this treatment modality safely in patients, even though our study demonstrated successful results in the treatment of implant-related chronic osteomyelitis in rats. (c) 2014 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 32:752-756, 2014.Öğe The measurement of alpha amanitin levels using hplc method in Amanita phalloides from Düzce province(Duzce University Medical School, 2012) Kaya, Ertuğrul; Karahan, Selim; Hancı, Mustafa; Yaykaşlı, Kürşat Oğuz; Sarıtaş, Ayhan; Bayram, Recep; Arslan, Seyfullah OktayAim: The aim of this study is to measure the level of alpha amanitin toxin using HPLC method from Amanita phalloides mushroom collected in the province of Düzce in 2010. Method: One of the mushrooms as a whole body. The other one was extracted after seperated into parts. The measurement was done by HPLC using 303 nm UV wavelength and 250x4,6 mm C18 5 ?m particle included column. Ammonium acetate+methanol+acetonitrile (80+10+10, v/v/v) was used as a mobile phase, and the flow rate was set 1 ml/min. The results were given as a toxin quantity in 1 g dry mushroom. Results: The amount of alpha amanitin was measured as 4,806 mg (±0,033) in the whole body, 3,522 mg (±0,024) in the cap, 5,318 mg (±0,056) in the lamellar, 0,903 mg (±0,004) in the ring, 2,577 mg (±0,037) in the stipe, 0,698 mg (±0,008) in the volva. Conclusion: The level of alpha amanitin in Amanita phalloides from Duzce Province is differ from different countries. Higher and lower levels of toxin than our data obtained investigations are present in the literature. The reason of this differences might be several factors like extraction methods, analysis methods and environmental conditions. © 2012 Düzce Medical Journal.Öğe Purification and in vitro toxicity of gamma amanitin(Taylor & Francis Ltd, 2015) Bakırcı, Sinan; Bayram, Recep; Yılmaz, İsmail; Yaykaşlı, Kürşat Oğuz; Bayram, Sait; Kaya, ErtuğrulWe aimed to obtain gamma amanitin with high purity through a purification process and compare toxic effects of alpha, beta, and gamma amanitin. Specific concentrations of the toxins (25, 10, 1, and 0.1 mu g/mL) were applied to the C3A human hepatocytes. A MTT test was performed to determine the level of toxicity. Alpha amanitin showed a higher toxicity in 48 h while the lowest toxicity was observed in beta amanitin. The toxicity level of gamma amanitin was found between the alpha and beta amanitin toxicity. Our method is suitable for obtaining gamma amanitin with high purity (>99%) as well as for obtaining alpha amanitin and beta amanitin. Gamma amanitin has been shown to have equal responsibility for toxicity as alpha amanitin in amanita poisoning.Öğe Toxin content and toxicological significance in different tissues and development stages of Lepiota brunneoincarnata mushroom(Taylor & Francis Ltd, 2015) Yılmaz, İsmail; Bakırcı, Sinan; Akata, Ilgaz; Bayram, Recep; Kaya, ErtuğrulWe investigated the quantity and concentration of toxins in different parts and in different growth phases of Lepiota brunneoincarnata mushroom. The amatoxins and phallotoxin levels were measured using the reversed phase high-performance liquid chromatography (RP-HPLC) method. Alpha amanitin (2.38 +/- 0.70mg/g) was followed by beta amanitin (1.97 +/- 0.52mg/g) and gamma amanitin (0.04 +/- 0.01mg/g) in trace amounts; it did not contain any phallotoxin. The cap part is richer in amatoxins than the stipe part. While medium mushrooms were quite rich in amatoxins, less levels of toxin were measured in fully developed mushrooms. The study showed in detail the toxin concentrations of L. brunneoincarnata with regard to different developmental stages and different segments.Öğe What is the restoring role of chrysin in alpha amanitin toxicity(A. CARBONE Editore, 2015) Bakırcı, Sinan; Bayram, Recep; Kaya, Ertuğrul; Yaykaşlı, Kürşat Oğuz; Yılmaz, İsmailAim: To explore whether chrysin has any protective effect against the deadly effect of alpha amanitin on hepatocytes that is one of the major toxins in the structure of Amanita phalloides, which is considered the most important mushroom responsible for fatal mushroom poisonings. Materials and methods: For this study, alpha amanitin was purified from A phalloides mushroom using the preparative HPLC (high performance liquid chromatography) method as described in the literature. Four hours after administering alpha amanitin in a concentration of 10 ?g/mL on the cells in a hepatocyte cell line (C3A), silibinin and chrysin were administered in various concentrations. The MTT [3-(4,5-Dimethylthiazol-2-yl)-2,5-Diphenyltetrazolium Bromide] test was used to determine cell viability. Results: The alpha amanitin was obtained in high purity from the mushrooms and was found to decrease cell viability down to a level of 63% after 48 hours due to its toxic effect on the liver cells in the cell culture. In the groups given silibinin and chrysin, both substances were seen to reduce the toxicity caused by alpha amanitin. Conclusion: Chrysin may play a role in decreasing the tissue damage caused by toxins and lowering the mortality rates in fatal mushroom poisonings associated with alpha amanitin.Öğe WHAT IS THE RESTORING ROLE OF CHRYSIN IN ALPHA AMANITIN TOXICITY?(Carbone Editore, 2015) Bakırcı, Sinan; Bayram, Recep; Kaya, Ertuğrul; Yaykaşlı, Kürşat Oğuz; Yılmaz, İsmailAim: To explore whether chrysin has any protective effect against the deadly effect of alpha amanitin on hepatocytes that is one of the major toxins in the structure of Amanita phalloides, which is considered the most important mushroom responsible for fatal mushroom poisonings. Materials and methods: For this study, alpha amanitin was purified from A phalloides mushroom using the preparative HPLC (high performance liquid chromatography) method as described in the literature. Four hours after administering alpha amanitin in a concentration of 10 mu g/mL on the cells in a hepatocyte cell line (C3A), silibinin and chrysin were administered in various concentrations. The MTT [3-(4,5-Dimethylthiazol-2-yl)-2,5-Diphenyltetrazolium Bromide] test was used to determine cell viability. Results: The alpha amanitin was obtained in high purity from the mushrooms and was found to decrease cell viability down to a level of 63% after 48 hours due to its toxic effect on the liver cells in the cell culture. In the groups given silibinin and chrysin, both substances were seen to reduce the toxicity caused by alpha amanitin. Conclusion: Chrysin may play a role in decreasing the tissue damage caused by toxins and lowering the mortality rates in fatal mushroom poisonings associated with alpha amanitin.Öğe Yüksek Saflıkta Beta Amanitin Üretimi(2012) Kaya, Ertuğrul; Yaykaşlı, Kürşat Oğuz; Karahan, Selim; Bayram, Recep; Sarıtaş, Ayhan; Yaykaşlı, EmineAmaç: Beta amanitin nadiren bilimsel araştırmalarda kullanılmakta ve kullanımı gün geçtikçe artmaktadır. Piyasada bu ürün %90 saflıkta ticari olarak satılmaktadır. Bu araştırmada yüksek saflıkta beta amanitin üretme yöntemi tanımlanmıştır. Yöntem: Saflaştırma işlemi Amanita phalloides mantarlarından ekstraksiyonla yapılmıştır. Öncelikle bu mantarlar toplanmış, ekstrakte edilmiş, 2 defa preparatif HPLC ile saflaştırma işlemi uygulanmıştır. Toksinin karşılaştırması, analitik HPLC sisteminde tutulma zamanı ve ultraviyole spektrumu karşılaştırılması yöntemleriyle yapılmıştır. Bulgular: İlk saflaştırma sonucunda elde edilen beta amanitin saflık oranı %91(2,36) olarak ölçülmüştür. İkinci saflaştırma sonucunda elde edilen beta amanitin saflık oranı %99,2(0,38) olarak ölçülmüştür. Saflaştırma sonucu elde ettiğimiz toksin ile beta amanitin standardın UV spektrumlarında her ikisinde de 303 nmde maksimum, 263 nmde minimum absorbans verdiği ve spektrum yapısının aynı olduğu görülmüştür. Sonuç: Tanımladığımız bu yöntemle, %99 saflıkta beta amanitinin üretilmesi mümkündür.
