Agaritine purified from Agaricus blazei Murrill exerts anti-tumor activity against leukemic cells.

BACKGROUND: Mushrooms of the genus Agaricus are a common folk remedy against carcinoma. The active ingredients, polysaccharides and protein-polysaccharide complexes containing beta-glucan, have been isolated and shown to have indirect tumor-suppressing activity via an immunological activation. METHODS: The diffusible fraction of a hot-water extract of Agaricus blazei Murrill (ABM) powder was fractionated by HPLC based on the anti-tumor activity against leukemic cells in vitro. The structure of the anti-tumor substance was determined by NMR and MS analyses. RESULTS: We purified a tumorcidal substance from the diffusible fraction of ABM and identified it as agaritine, beta-N-(gamma-l(+)-glutamyl)-4-(hydroxymethyl) phenylhydrazine, having a molecular mass of 267 Da. This compound inhibited the proliferation of leukemic cell lines such as U937, MOLT4, HL60 and K562 with IC(50) values of 2.7, 9.4, 13.0, and 16.0 microg/mL, respectively, but showed no significant effect on normal lymphatic cells at concentrations up to 40 microg/mL. Although agaritine has been suspected of having genotoxic or carcinogenic properties, agaritine did not activate the umu gene of Salmonella, which reacts to carcinogens. GENERAL SIGNIFICANCE: The results indicate that agaritine from ABM has direct anti-tumor activity against leukemic tumor cells in vitro. This is in contrast to the carcinogenic activity previously ascribed to this compound. Our results also show that this activity is distinct from that of beta-glucan, which indirectly suppresses proliferation of tumor cells.

Identification and significance of phenazone drugs and their metabolites in ground- and drinking water.

Residues of three phenazone-type pharmaceuticals have been identified in routine analyses of groundwater samples from selected areas in the north-western districts of Berlin, Germany. Phenazone, propiphenazone, and dimethylaminophenazone have been detected in some wells at concentrations up to the low microg/l-level. Additionally, three phenazone-type metabolites namely 1-acetyl-1-methyl-2-dimethyl-oxamoyl-2-phenylhydrazide (AMDOPH), 1-acetyl-1-methyl-2-phenylhydrazide, and dimethyloxalamide acid-(N'-methyl-N-phenyl)-hydrazide have also been identified in these groundwater samples. The residues are suspected to originate from former production spills of a pharmaceutical plant located in a city north of Berlin. It was observed that with the exception of AMDOPH all other residues were efficiently removed during conventional drinking water treatment. The drug metabolite AMDOPH deriving from dimethylaminophenazone residues was found at concentrations of 0.9 microg/l in finished drinking water. However, a following study on the toxicological relevance of the AMDOPH residues has shown that there is no toxicological harm for humans at the low concentrations of AMDOPH observed in Berlin drinking water.

Agaritine does not mediate the mutagenicity of the edible mushroom Agaricus bisporus.

Ethanolic extracts of the edible mushroom Agaricus bisporus displayed a direct-acting mutagenic response in various Salmonella typhimurium strains, TA104 being clearly the most sensitive. Incorporation of an activation system derived from the liver of mice, hamsters or Aroclor 1254-induced rats failed to increase the mutagenic response. The mutagenic response of ethanolic extracts from various types of mushroom, containing different levels of agaritine (range 0.3-6.5 g/kg fresh weight), was very similar and did not correlate with the agaritine levels. Moreover, use of gamma-glutamyl transpeptidase, the enzyme catalysing the activation of agaritine, as an activation system did not enhance the mutagenicity of the mushroom ethanolic extracts. It is concluded that agaritine is not responsible for the mutagenicity of mushroom extracts.

Occurrence, stability and decomposition of beta-N [gamma-L(+)-glutamyl]-4-hydroxymethylphenylhydrazine (agaritine) from the mushroom Agaricus bisporus.

A chromatographic technique was developed that could clearly separate beta-N [gamma-L(+)-glutamyl]-4-hydroxymethylphenylhydrazine (agaritine) from all other components in 10-500-microliters samples of mushroom extracts. Locally purchased mushrooms were found to contain mean levels of 0.4 - 0.7 mg agaritine g. The agaritine content of the mushrooms had decreased by 2-47% after 1 wk of storage in a domestic refrigerator and by 36-76% after 2 wk of such storage. Canned mushroom soup and canned mushrooms did not contain detectable agaritine; a sample of frozen mushrooms contained a mean level of 0.33 mg/g and a batch of fresh mushrooms lost about 32% of their agaritine content on cooking. In mice given 3 mg agaritine by gavage, agaritine was detected in all parts of the gastro-intestinal tract 15 min after dosing, but none was detectable in the gut after 3 hr. The enzyme gamma-glutamyltranspeptidase derived from pig's kidney was found to be capable of decomposing agaritine to glutamic acid and 4-(hydroxymethyl)phenylhydrazine, and to have nine times such activity as an enzyme isolated from mushrooms.