Further Studies on the Fractionation of Human Pituitary Glands1

Parlow, Albert F.; Wilhelmi, Alfred E.; Reichert, Leo E.

doi:10.1210/endo-77-6-1126

Cited by 48 publications

(25 citation statements)

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“…At the same time, more than 95 % of the HGH and probably a similar part of the FSH were unadsorbed. The statements concerning separation of FSH from LH at this step are based not only on the data in Table 2 and later in this paper but on previous publications of ourselves and others (5,(16)(17)(18)(19)(20). That this step was such a good method for removing growth hormone from the TSH-LH mixture has not been shown previously.…”

Section: Volume-mentioning

confidence: 64%

Further Studies on the Purification of Human Thyrotropin

Bates

Garrison

Cooper³

et al. 1968

Endocrinology

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Thyrotropin activity was concentrated 1400-fold from acetone dried human pituitary glands. The final product had a TSH potency of 10.4 Human International Units/mg. The yield of activity was 34 %. The glands were extracted by percolation in stepwise sequence with 76% ethanol-2% NaCl, 50% ethanol-5% NaCl and water. The 50% ethanol-5% NaCl solvent front contained most of the TSH, LH and FSH. TSH and LH were separated from FSH and most of the contaminating GH by adsorption of the TSH and LH at pH 6 on carboxymethyl cellulose. The TSH was then separated from the LH by adsorbing the TSH on diethylaminoethyl cellulose at pH 9.5. An LH fraction with a potency of 5.5 mg equivalents of NIH-LH-S-1/mg was obtained that had a TSH activity of only 0.01 Human IU/mg. This LH fraction was 920 times more potent than the acetone dried glands. By the above procedure the yields of TSH were superior, of FSH and LH were the same, and of GH and prolactin were inferior to those of other laboratories that used more aqueous methods of extraction. {Endocri-nology 83: 721, 1968) P ERHAPS the greatest unsolved problem in isolation of the hormones from the anterior pituitary is how to extract in a quantitative manner all of the hormones simultaneously. To complicate matters further, the various hormones show species differences in extractability, stability and solubility with various solvents. For example, the glacial acetic acid extraction method (1) is excellent for growth hormone and ACTH but destroys most of the other hormones, and ACTH rapidly disappears during neutral or alkaline extraction in aqueous media (2). With the aim of recovering all hormones, Wilhelmi (3) has devised a sequence of extraction and fractionation which is satisfactory in general except for low recovery of thyrotropin (TSH). Our percolation or "dry to wet" extraction procedure starting with 95% ethanol (4) has been developed with the aim of preventing enzyme action and of recovering all hormones. This goal of quantitative recovery of all hormones has not been en- Received February 2, 1968. tirely achieved but the method has given excellent yields of TSH from glands (5, 6) or plasma (7) and, as shown below, of FSH and LH from the current batch of human glands. Saffran, Muhlstock and Caplan have extracted ACTH by percolation (8). This batch of human pituitary glands was processed, at the request of the National Pituitary Agency, with the primary aim of purification and isolation of human TSH in sufficient quantity and purity that it would be suitable for distribution and use in radioimmunoassay. Materials and MethodsAbout 10,000 human pituitary glands that had been collected under acetone were received from the National Pituitary Agency, Baltimore, Md. They were dried in air. Approximately 10% of the glands were white. These were picked out and weighed 73 g after milling. Bioassays for TSH showed them to have less than half the potency of the tan colored glands; hence they were not extracted. The tan colored glands were milled through a 40 mesh screen in a Wiley m...

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Section: Volume-mentioning

confidence: 64%

Further Studies on the Purification of Human Thyrotropin

Bates

Garrison

Cooper³

et al. 1968

Endocrinology

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“…Human plasmin was purchased from KABI, Stockholm, Sweden. Purified hGH was prepared as described previously (18,19). Most preparations of hGH contain multiple active components (20).…”

Section: Methodsmentioning

confidence: 99%

“…was measured by the 9-day weight gain test performed as previously described (18 (24). After incubation, the cartilages were boiled for 5 min and washed overnight in running water.…”

Section: Methodsmentioning

confidence: 99%

Metabolic Effects of Plasmin Digests of Human Growth Hormone in the Rat and Man

Mills¹,

Reagan²,

Rudman³

et al. 1973

J. Clin. Invest.

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A B S T R A C T As a first step in our study of structurefunction relationships among primate and non-primate growth hormones, human growth hormone (hGH) was subjected to the limited digestive activity of human plasmin. The lyophilized whole digest, containing less than 2% of unchanged hormone, had an average of 2. The digest also caused glucosuria in partially pancreatectomized rats treated with dexamethasone.These metabolic actions of plasmin-digested hGH in the array of animal tests were confirmed by comparable effects elicited in 11 human subjects (nine pituitary-deficient children and adolescents and two nondeficient adults). A single injection of the plasmin digest caused an increase in plasma free fatty acids and a fall in plasma amino acids. Seven daily injections caused positive balances of nitrogen, phosphorous, sodium, and potassium, gain in body weight, and in two of three subjects impairment of glucose tolerance. The potency of the plasmin digest in producing these metabolic effects in man was comparable to that of native hGH.Thus, 2-3 bonds in the hGH molecule can be cleaved by plasmin without impairing the hormone's growth-

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“…The three schedules were said to be generally equipotent, but data supporting this conclusion were not presented. In the hypophysectomized rat, however, injection of half a specified dose of bovine GH twice a day gave a greater response than injection of the entire dose once a day in the morning (23).…”

mentioning

confidence: 87%

Diurnal Variation in the Responsiveness of Human Subjects to Human Growth Hormone

Rudman¹,

Freides²,

Patterson³

et al. 1973

J. Clin. Invest.

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A B S T R A C T The objective of this study was to compare the responsiveness of human subjects to the anabolic effects of human growth hormone (HGH) administered at 8 a.m. or at 11 p.m. Three doses of HGH were used: A, 0.0168 U/kg body weight (BW)"'4 per day; B, 0.0532 U/kg BW"/' per day; C, 0.168 U/kg BW"'4 per day. The effect of each dose on daily balances of N, P, Na, and K and on BW was measured. The subjects were of two groups: (a) seven GH-deficient children, of whom three were deficient in ACTH; and (b) three patients with limb-girdle dystrophy. ACTHdeficient patients in group (a) received exogenous cortisol at 7 a.m. In all 10 subjects, the anabolic effects of dose C, and sometimes of B and A, administered at 11 p.m. were significantly (P < 0.05) greater than when administered at 8 a.m. In these experiments plasma cortisol concentration averaged 3 times greater at 8 a.m. than at 11 p.m.In the next experiments, exogenous cortisol was administered to the three ACTH-deficient patients at 10 p.m. and the responsiveness to HGH injected at 11 p.m. vs. 8 a.m. was again compared. Under these conditions, when plasma cortisol concentration averaged 3 times greater at 11 p.m. than at 8 a.m., HGH injected at 8 a.m. caused significantly greater anabolic responses than HGH injected at 11 p.m.These findings indicate that the magnitude of the anabolic response to exogenous HGH is inversely related to the plasma cortisol concentration at the time of HGH injection.

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Further Studies on the Fractionation of Human Pituitary Glands1

Cited by 48 publications

References 0 publications

Further Studies on the Purification of Human Thyrotropin

Further Studies on the Purification of Human Thyrotropin

Metabolic Effects of Plasmin Digests of Human Growth Hormone in the Rat and Man

Diurnal Variation in the Responsiveness of Human Subjects to Human Growth Hormone

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