Normal human urine contains methanol largely as methoxylated compounds; a small quantity of free methanol is also present. The methoxylated compounds exist partly as ether-soluble compounds, for example methylated phenolic acids, and partly as methoxylated compounds not extracted by ether from acid solution. The latter may be related chemically to quercitrin. Feeding experiments suggest that 3,4dihydroxyphenolic substances are largely methylated while with other phenolic substances methylation is minimal. THE present communication is an extension of previous studies.'?*. It has been shown that, in man tannic and 3,4-dihydroxybenzoic acids are largely methylated to 4-hydroxy-3-methoxy derivatives. The present paper is concerned with an examination of the nature, in very general terms, of methoxy compounds in human urine and also of the extent to which phenols and phenolic acids, other than those with an ortho dihydroxy structure, are methylated. METHODSThe general principles of the techniques employed have been described2. The following fractions have been determined.( a ) Free methanol. 10 ml. of urine diluted to 15 ml. with water was heated to boiling in a 100 ml. R.B. flask attached to a water cooled condenser and 10 ml. of distillate collected.( b ) Methanol liberated by the action of hot strong sulphuric acid on methoxylated compounds present in untreated urine. Into a 100 ml. R.B. flask attached to a water cooled condenser (all glass equipment) were introduced 10 ml. of urine and 5 ml. of concentrated sulphuric acid. The mixture was heated to boiling and the distillate collected. Heating was continued until the sulphuric acid reached the fuming stage after which the mixture was allowed to cool. After the addition of 5 ml. of water, the mixture was again heated, the sulphuric acid being allowed to reach the fuming stage. This part of the procedure was repeated, so that three distillates in all were collected. A knife point of sodium bicarbonate was added to the combined distillates. The mixture was heated in apparatus similar to that described above and 10 ml. of distillate collected.(c) Methanol liberated by the action of hot strong sulphuric acid on the acidic fraction of urine. 10 ml. of urine and 1 ml. of 1 0~ hydrochloric acid in a test tube were heated in a boiling water bath for 1 hour. After cooling, the urine was extracted three times with 40 ml. quantities of redistilled ether. The combined ether extracts were evaporated to dryness. 62

The Mitchell reaction (ferrous sulphate) has been applied to the determination of catecholic phenolic acids and pyrogallol in urine. A technique has been described for the determination of pyrogallol in the presence of catecholic phenolic acids. The separation of catecholic phenolic acids by paper chromatography has been examined. A study has been made of the excretion of these substances in urine.

A method has been described for the determination of the methoxy group in phenolic compounds. Tannic and 3 : 4-dihydroxybenzoic acids when administered orally to man are excreted in appreciable quantities as 4-hydroxy 3-methoxybenzoic and m-hydroxybenzoic acids.THE present communication is an extension of the studies reported previously1. Human urine contains appreciable quantities of m-hydroxyand 4-hydroxy-3-methoxyphenolic a c i d F . The origin of the m-hydroxy acids has been obscure since they do not occur in nature nor is their source dietary. Paper chromatographic studies suggest that 3 : 4-dihydroxyphenolic derivatives may be dehydroxylated to produce mhydroxyphenyl compounds and also methylated to produce 4-hydroxy 3 -m e t h o~y p h e n y l~~~~~J~.The present paper is concerned with quantitative studies involving the determination of the methoxy group and the colorimetric determination of m-hydroxybenzoic and vanillic acids.

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A procedure for the identification of the synthetic oestrogens, p‐hydroxypheno‐barbitone and 5‐(p‐hydroxyphenyl) 5‐phenylhydantoin in urine by use of two dimensional paper chromatography is described.