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The incorporation of radioactivity into total protein, albumin-like protein and albumin in liver, and into total protein and albumin in blood plasma, was studied for a time period of 2 h after intraportal injection of ~-[l-'~C]leucine.1. In the liver, radioactivity began to increase in albumin-like protein before 2.5 min and in albumin at about 10 min after injection. In the plasma, radioactive albumin appeared at about 15 min. No albumin-like protein could be detected in the plasma.2. Maximum radioactivity was reached first in albumin-like protein, then in hepatic albumin and finally in albumin in the blood-stream. The maximum specific radioactivity of albumin-like protein was 15 times higher than that of extravascular hepatic albumin which, in turn, was 6 times higher than that of plasma albumin.3. The increase of radioactivity in albumin in the blood corresponded almost quantitatively to the decrease of radioactivity in albumin-like protein.4. It is concluded that the albumin-like protein is the precursor of albumin in vivo. It is converted into albumin 5 -6 min before the appearance of newly synthesized albumin in the blood-stream.In contrast to the easy isolation from serum, rather extensive purification is required to obtain pure albumin from biological material involved in albumin synthesis, such as liver [l-41, hepatoma [2,3], hepatocyte suspensions [ 5 ] , and cell-free proteinsynthesizing systems from liver [6-81. This is due to the presence in these tissues of a protein similar to albumin [6,7,9,10]. When applying appropriate purification procedures, Judah and Nicholls observed that liver slices continued to incorporate ['4C]leucine into albumin after incorporation into total protein had been interrupted by cycloheximide or excess unlabelled leucine [Ill. This suggested that a precursor protein other than intrahepatic albumin is involved in the biosynthesis of serum albumin. Recently, a possible precursor protein, an albumin-like protein, was isolated from liver and characterised [12-141. It differed from albumin by a short peptide extension at the N-terminus. The earlier finding of identical Ntermini for albumin-like protein and albumin [15] can probably be explained by insufficient separation of the two proteins and difficulties in the determination of the N-terminal amino acid of the albumin-like protein.In this paper, we describe the kinetics in vivo of ['4C]leucine incorporation into total protein, albuminlike protein and albumin in the liver, and into total protein and albumin in the plasma. A quantitative analysis provides evidence that the albumin-like protein is a precursor in the synthesis of albumin in vivo. A preliminary report of this work appeared elsewhere P61. MATERIALS AND METHODS AnimalsInbred buffalo rats were kept at a room temperature of 23 "C and had free access to water and a diet containing about 20 % protein (Barastoc Mouse Breeder Ration, Barastoc Products, Melbourne, Australia). Only male rats were used. They were starved overnight and operated on between 9 a.m. and 2 p.m. ...

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The sequence of radioactively labelled amino acids at the N‐terminus of proalbumin was determined by automated Edman‐degradation. [3H] Valine, [3H] phenylalanine or [14Carginine was incorporated into protein in vivo for a time period of 10 min after injection. Since albumin remains unlabelled during this time period (Urban et al., 1976), separation of proalbumin and albumin was not required for this work. Hence, compared to previous methods, a shorter purification procedure could be used which increased the yield of anti‐albumin‐precipitable protein and reduced the risk of proteolysis. Microsomes were prepared from livers removed 10 min after injection of the radioactively labelled amino acids. A buffer extract of the acetone‐dried powder from these microsomes was chromatographed on DEAE‐cellulose. All protein obtained after chromatography which could be precipitated with antiserum to serum albumin was isolated by immunoprecipitation and subsequent separation of the antigen‐antibody complex. The sequence of radioactive amino acids in this antigen preparation suggests that about 20–25% of proalbumin possessed at the N‐terminus the pentapeptide sequence X‐Val‐Phe‐Arg‐Arg‐ whereas 75–80% contained the hexapeptide sequence Arg‐X‐Val‐Phe‐Arg‐Arg‐.