A new method for the fractionation of dough made from wheat flour is described. The method does not involve the use of diluents or extractants and consists simply of ultracentrifugation of the dough. With this method dough is resolved into from five to seven discrete layers, depending on the dough treatment. A study of the fractionation of doughs made from seven Australian flours is described together with the physical and chemical composition of the various layers.

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SummaryCrude tetanus toxin and toxoid were prepared by methanol precipitation. The toxin was purified by a combination of TEAE·cellulose and Sephadex G·200 chromatography at pH values less than 6· o. The toxoid was purified by DEAE· cellulose at approximately neutral pH values. The nature and amount of amino acids of the culture medium which had condensed with the tetanus toxoid proteins during detoxification with formaldehyde was determined.

Tetanus toxin has been extracted from culture filtrates by a single, cold methanol precipitation, followed by gradient elution on DEAE-cellulose. Immunoelectrophoretic evidence indicated that most of the bacterial protein was precipitated by the methanol. Selection of the toxin elution peak gave a product having only one immunoelectrophoresis precipitin line yet containing 50--60% of the Lf units applied to the column. The toxicity of this product was comparable to that found by other workers, using other methods of preparation. Sedimentation values of the concentrated pure toxin indicated heterogeneity in molecular aggregation and under conditions used no 4 S material could be found. Amino acid analysis of the tetanus protein is also reported.

SummaryDoughs made from Australian wheat flours were fractionated by ultracentrifugation. The disulphide and sulphydryl contents of the fractions were determined polarographically. A low molecular weight, sulphydryl-containing fraction was found in doughs made from poor quality flours.Up to 26% of the sulphydryl groups originally present in the flours disappeared during dough formation. Doughs mixed in the presence of air or iodate showed a rapid initial, and subsequent more gradual loss of sulphydryl groups as mixing progressed, or as the concentration of iodate increased. The sulphydryl groups of the soluble components of dough were more labile during overmixing, and in the presence of iodate and N -ethyl maleimide than were the sulphydryl groups of the gluten complex.

Summary[S6S]Cysteine was added, during mixing, to doughs made from wheat flour, in amounts which did not significantly affect the level of endogenous diffusible sulphydryl compounds in flour and which produced no change in the rheological properties of the dough. The doughs were fractionated by ultracentrifugation and analysis of the distribution of the isotope in the dough fractions demonstrated that [S6S]cysteine was bound to both soluble and gluten proteins by disulphide bonds.[14C]Leucine did not become bound to the fractions of dough under the same conditions. Incorporation of cysteine into dough was increased in the a.bsence of air, suggesting disulphide--sulphydryl exchange rather than oxidation as the mode of incorporation. The amount of isotope bound to protein was also increased either by lengthening the times of mixing or relaxation of the dough.The incorporated [S6S]cysteine could be released in dough only by the addition of large excess of unlabelled cysteine.

Chicken erythrocyte .a.histone, whose behaviour on moving boundary electro-phoresis or on ultracentrifugation did not indicate any heterogeneity, has been resolved into seven components by starch.gel electrophoresis.

The binding of formaldehyde with tetanus toxin under mild detoxificatIOn conditions (0 �1M phosphate buffer, pH 7� 0, O� 005-0� 06M HCHO, 20-37�C) has been studied quantitatively by using [14C]formaldehyde. When detoxification was followed with time (0� 005M HCHO, 37�C) the toxin showed a characteristic immediate drop in lethality (~80%), followed by a slower detoxification, whereas the combination of the HCHO was a more gradual process. When a fixed time period was used (5 days, 20 and 37�C) at least O' 02M HCHO was required to destroy the lethality of the toxin towards mice, and at this concentration approximately 60 moles of HCHO were bound per mole of toxin. Fractionation of a partially detoxified toxin indicated the presence of a range of molecular species with varying formaldehyde binding and lethality.