EDIBLE MUSHROOMS AS FUNCTIONAL FOODS Professor Theeshan BAHORUN - PowerPoint PPT Presentation

EDIBLE MUSHROOMS AS FUNCTIONAL FOODS Professor Theeshan BAHORUN National Research Chair (Mauritius Research Council) ANDI Center for Biomedical and Biomaterials Research University of Mauritiu s

Mushrooms Mushrooms as High in Protein (mean functional food proportion of 33.4%) Low in fats Calories (2-6% dry matter) (20.3-74.3%) Relatively rich in riboflavin, niacin, ergocalciferol Elevated levels of phosphorus and potassium and fairly high contents of magnesium ERGOTHIONEINE PHENOLIC Antitumor, Main Classes a potential ACIDS antibacterial, of Phenolic antioxidant present and antifungal Compounds in most mushroom properties FLAVONOIDS species

Anticancer Properties of Mushrooms Clinical trials in cancer patients By countering the side- Complement effects of cancer e.g. Chemopreventive and chemotherapy and nausea, bone marrow antitumour activities radiation therapy suppression, anemia, and lowered resistance Ganoderma lucidum “Mushroom of Immortality” Research in edible mushrooms is gaining popularity In vitro and in vivo anticancer activities of A garicus bisporus Possess Proteoglycans derived Suppress aromatase activity immuno- from fruiting body and and estrogen biosynthesis modulatory and mycelia of Pleurotus antitumor ostreatus properties Potential use in treatment of breast cancer

Mushroom Samples Agaricus bisporus Pleurotus sp. CC 201 CC 200 Pleurotus sajor-caju Pleurotus Hybrids Commercially available Mushroom Unit (S.K.C Surat and Co Ltd) (Food and Agricultural Research and Extension Unit) Obtained at optimum maturity Harvested at optimum maturity stage stage Methanolic extracts obtained by exhaustive extraction Total Proanthocyanidin HPLC Analysis Total Phenolic content Total Flavonoid content content

Phenolic Profile of Mushrooms  Total Phenolic content of the mushroom extracts ranged from 33.28 to 133.69 mg/g DW. Highest level of Maximum total phenolics flavonoid levels (133.69 ± 3.204 (4.63 ± 0.052 mg/g mg/g DW) DW) Negligible Negligible proanthocyanidins proanthocyanidins

Phenolic Profile of Mushrooms HPLC Analysis of phenolic compounds and ergothioneine in the four mushroom extracts Pleurotus sajor- Pleurotus hybrid Pleurotus hybrid Agaricus caju (CC 114) (CC 201) (CC 200) bisporus Phenolic content of mushrooms (µg/g DW) I Gallic acid 356.90 b ± 38.922 240.34 c ± 12.186 224.71 c ± 4.364 726.17 a ± 4.263 II Protocatechuic 630.00 a ± 15.161 590.52 b ± 15.267 362.91 c ± 10.469 84.85 d ± 16.143 acid IV Pyrogallol 2831.32 c ± 5457.03 b ± 7791.24 a ± 2354.68 d ± 105.838 149.911 121.08 88.626 Ergothioneine content of mushrooms (µg/g DW) III Ergothioneine 2518.88 a ± 1596.867 c ± 1351.04 d ± 2261.18 b ± 22.181 13.958 13.116 14.563 DW: Dry weight; I Retention time (R.T)=5.787; II R.T=11.773; III R.T=3.829; IV R.T=7.339; Data expressed as mean ± standard deviation (n=5); ANOVA and Fisher’s LSD Test at 5% significance level; Common superscripts between rows represent no significant difference between mushroom samples.

Animal Study Balb/c mice (7 weeks of age, 20 ± 2 g, n=120) ME 2 ME 1 Group II- Carcinogen Group I-Control treated Treatment Pleurotus sajor-caju Agaricus bisporus Period of 3 months Group Dose (mg/kg b.w.) Group Dose (mg/kg b.w.) Phosphate Saline N-methyl, N- VIII 150 + MNU Buffer (PBS) only nitrosourea (MNU) III 150 + MNU (i.p.) at 50 mg/kg body (i.p.) IX 300 + MNU weight (i.p.) IV 300 + MNU (i.p.) (i.p.) X 450 + MNU Mice Liver V 450 + MNU (i.p.) Blood Sample DNA (i.p.) Histological XI 600 + MNU Analyses VI 600 + MNU (i.p.) Haematological (i.p.) XII 600 Laser Raman Biochemical VII 600 Spectroscopy Analyses

Diets high in nitrostable foods cause cancer  high levels of nitrates used in food • Examples of foods naturally high in nitrates: preservation are carcinogenic Fish, oysters, mussels, crab, lobster, Chinese cabbage, some leafy vegetables, Cigarette smoke, Beer & wine, Cheese, luncheon & sausage meats, Canned foods A possible reason why Asians have high rates of stomach & mouth cancers? Example: N-methyl-N-nitrosurea (MNU) MNU causes several cancers in animal models. It targets the liver in mice Methyl group transferred to nucleobases Causes AT:GC mutations

The liver/body weight ratio is highly indicative of tumour presence Liver:Body Weight Ratio(%) d XII-ME2 only (600 mg/kg) c XI-ME2 (600 mg/Kg) + MNU X-ME2 (450 mg/Kg) + MNU c IX-ME2 (300 mg/Kg) + MNU d c VIII-ME2 (150 mg/Kg) + MNU c VII-ME1 only (600 mg/kg) b VI-ME1 (600 mg/Kg) + MNU V-ME1 (450 mg/Kg) + MNU c 2.8 folds increase IV-ME1 (300 mg/Kg) + MNU b in L:BW ratio III-ME1 (150 mg/Kg) + MNU c a II-MNU only I-Normal (PBS only) d 0 5 10 15 Liver:Body weight ratio for the 12 treatment groups Data expressed as mean + standard deviation (error bars) (n=5); ANOVA and LSD at 5% significance; Similar superscripts on the mean values represent no significant differences between the treatment groups.

• Smooth PBS-treated mice and even growth of hairs in normal mice • Healthy liver A B C • Pachy skin and uneven MNU-treated mice shedding of hairs • H&E stained liver section: Nucleo- cytoplasmic ratio (cells appearing leaky), D E F enlarged vacuolated hepatocytes, altered cell MNU + ME 1 300mg/kg structure Extract Protective effect: G H I • Reduction in hair loss • Reduction in lesions MNU + ME 2 450mg/kg • Cell architecture almost comparative to PBS J K L Morphological changes in mice and liver, and H&E stained liver sections from PBS, MNU, MNU+ME 1 300mg/Kg and ME 2 450mg/Kg groups after 3 months supplementation.

Reduction in lesions and mushroom protective effects (MNU + 300 mg/kg Mushroom A e xtract) Healthy liver of Balb/c mice C MNU A (N-Methyl-N-nitrosourea) B Histopathology of liver tissue Micronodular lesions in liver of MNU-treated mice B A D ONGOING Reduction in lesions and mushroom protective effects Molecular Mechanisms Works (MNU + 450 mg/kg Mushroom B extract) University of Seoul, South Korea C D

Laser Raman Spectroscopy Continuous exposure to MNU N-methyl-N- nitrosourea Leads to an Increased AT:GC mutation formation, repair and persistence of DNA adducts Inflammation and  MNU acts as an alkylating agent reacting with elevated levels of nucleophillic nitrogen, oxygen atoms in bases and DNA inflammatory cytokines (IL-1b, IL- phosphate groups to create mutagenic lesions. 6)  The region 1200 – 1600 cm − 1 (assigned to purines and Creating a micro- pyramidines) corresponds to nucleic bases which are environment prone to any type of alkylation by MNU. conducive for the survival and development of  Mushroom protective effect was confirmed by Raman cancer cells. spectroscopy where, the MNU-DNA interaction as evidenced by an intense peak at 1254 cm -1 was normalised. and was not apparent in any of the mushroom-treated DNA samples.

Alkylation at adenine bases, resulting in a strong interaction between MNU and DNA structure .

Prospects USA: Commercial Production of Agaricus bisporus Cheap source of protein with Use of hallucinogenic ease of home mushrooms in Mesoamerica production 500 B.C. – 900 A.D. 2020 1880 2016 Scientific Data 300 edible mushroom species identified on Mauritian 20 fold increase in mushroom production mushrooms Medicinal mushrooms and their derivatives-Clinical trials Mauritius ? Scope for increasing cultivation Still not meeting Production of active extracts local demand, high imports

Acknowledgements Funding for Research Work – National Research Chair Program ( MRC) – Chhatrapati Shahu Ji Maharaj (CSJM) University (Kanpur, India) Animal and Raman Spectroscopy facilities – Institute of Biosciences and Biotechnology, CSJM University) – Department of Physics, Banaras Hindu University, (Varanasi, India)

Prof (Dr) Theeshan Bahorun Dr Vidushi Dr Shameem Neergheen-Bhujun Fawdar Mr Nawraj Ms Darshini Rummun Narrain Biopharmaceutical unit Ms Srishti Ms Piteesha Ramsaha Ramlagan Ms Rima Ms C. Tatsha Beesoo Bholah Mr Arvind Gopeechund