The simple code names adopted refer to the different substituents in the basic ring system, HH referring to the ring methylene, HMe to the ring methyl-methylene, Me to the methyls of the acetyl groups and OEt to the ethoxy groups of the ester side chains. See also Figs. 5 and 6. One recrystallization from ethanol was sufficient to give a product whose melting point agreed with that given in the literature, but up to three more were required before the colour of the crystals did not change further. There was also a marked increase in the fluorescence, particularly in the last compound (HMe/OEt). The melting points were also raised slightly, particularly the first compound (HH/Me, or DDL). These are given in Table 3, together with the analyses. As found by others (Hantzsch, Knoevenagel, quoted by Schiff & Prosio, 1895) there was a general deflciency in the carbon content. Repeat analyses on HH/Me gave figures for carbon of 66-7, 67-7 and 68-6% (averaged for Table 3), while two using a copper tube for combustion gave 69-4 and 69-7 %. Absorption spectra. It is clear from Figs. 5 and 6 that the compound HH/Me (DDL) is in no way exceptional. The strong-yellow colour is due to the absorption extending sufficiently far into the visible region; only the last com, pound, HMe/OEt, was finally obtained as completely colourless crystals, the two intermediate ones showing some absorption in the extreme blue. The optical properties are summarized in Table 2. SUMMARY A colour reaction for formaldehyde is described which depends on the synthesis of diacetyldihydrolutidine from acetylacetone and formaldehyde in the presence ofexcess of ammonium salt. The conditions are mild enough to allow of its use with living material, and its sensitivity and degree of specificity are comparable to those of other reactions requiring more severe conditions. The work arose from investigations carried out on behalf of the

THE following metabolic products, present in the metabolism solution of Penicillium griseo-fulvum Dierckx when this mould is grown under different cultural conditions and on different culture solutions, have been isolated in this laboratory: 6-hydroxy-2-methylbenzoic acid [Anslow & Raistrick, 1931], 2:5-dihydroxybenzoic acid (gentisic acid), fumaric acid and mannitol [Raistrick & Simonart, 1933] and fulvic acid, C14H1208, a yellow crystalline substance of at present undetermined molecular constitution [Oxford et al. 1935]. The purpose of the present communication is to record observations on a hitherto undescribed chlorine-containing metabolic product of P. griseo-fulvum which has been isolated from the mycelium of this mould and for which the name griseofulvin is proposed.Griseofulvin, C17H.706C, M.P. 218-219', is a colourless, crystalline, neutral compound giving no colour with FeCl3 and containing no free hydroxyl or carboxyl groups. It is highly dextrorotatory, [ac],61+ 417g.One oxygen atom is present as >CO since a crystalline mono-oxime, C,7H1806NC1, was readily obtained; derivatives are also formed with phenylhydrazine and with 2:4-dinitrophenylhydrazine, but these could not be obtained crystalline. The function of three other oxygen atoms is obvious since griseofulvin contains three methoxyl groups. A study of the products obtained on acid and alkaline hydrolysis showed that griseofulvin must be the methyl ester of a carboxylic acid, and hence the function of a fifth oxygen atom is accounted for. The function of the sixth oxygen atom has not been definitely settled though evidence given later indicates that it is present as an oxygen bridge.Griseofulvin, when hydrolysed with boiling N aqueous-alcoholic H2SO4, yields griseofulvic acid, C16H1506C1, [x]5461+ 5080, a monobasic acid containing two methoxyl groups and giving only a feeble colour with FeCl3, but these facts in themselves are not sufficient to establish the presence of the grouping -COO. CH3. However, hydrolysis of griseofulvin, or further hydrolysis of griseofulvic acid, with boiling aqueous N/2 NaOH yields norgriseofulvic acid, C15H130O6C1, [a]05M+609', a dibasic acid containing only one methoxyl group, together with decarboxygriseofulvic acid, C,5H1504C1, [x]e1 -310, an insoluble neutral compound containing two methoxyl groups, giving no colour with FeCl3, and derived from griseofulvic acid by the loss of 1 mol. of CO2. Decarboxygriseofulvic acid is stable to acid hydrolysis and hence it seems certain that griseofulvin contains only one -COO.COH3 group and that the second acidic ( 240 )

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