Isolation of two novel E prostaglandins in human seminal fluid

cis-8,11,14,17-[1-14C]Eicosatetraenoic acid was incubated with microsomes of ram seminal vesicles and 1 mM glutathione for 3 min at 37 degrees C. The main metabolite was identified as 17,18-dehydroprostaglandin E1 by capillary column gas chromatography-mass spectrometry. Human seminal fluid was analyzed for the presence of 17,18-dehydroprostaglandin E1 and prostaglandin E3. Whereas prostaglandin E3 could be demonstrated by capillary gas chromatography-mass spectrometry, 17,18-dehydroprostaglandin E1 could not be found under these conditions. However, human seminal fluid contained two compounds with a similar polarity on reversed phase high performance liquid chromatography as 17,18-dehydroprostaglandin E1 and prostaglandin E3. The two compounds were identified as 18,19-dehydroprostaglandin E1 and 18,19-dehydroprostaglandin E2 by gas chromatography-mass spectrometry, by UV analysis after conversion to the corresponding prostaglandin B compounds, and by ozonolysis. The amount of each of the two prostaglandins in human seminal fluid seemed to be in the same order of magnitude as the amount of prostaglandin E3.     

gamma-Glutamyl transpeptidase of human seminal fluid.

γ-Glutamyl transpeptidase, which is present in high levels in human seminal fluid plasma, was purified about 870-fold from this source. The enzyme is present in seminal fluid plasma in particulate form. Purification by a procedure involving treatment with bromelain gave a protein (apparent molecular weight, about 70,000), which exhibited catalytic properties characteristic of γ-glutamyl transpeptidase preparations isolated from rat kidney and other mammalian tissues. The physiological significance of seminal fluid γ-glutamyl transpeptidase and its potential clinical value are considered.     

Creatine kinase and ATPase in human seminal fluid and prostatic fluid

A creatine kinase assay based on estimation of creatine liberated from creatine phosphate was accurate and reproducible for use with seminal or prostatic fluid, after allowance was made for acid phosphatase interference. Comparison of this method with one which relies on enzymic coupling of ATP formation to NADP+ oxidation shows that the latter under-estimates creatine kinase activity by a factor of about 3. This discrepancy could be due to the high ATPase activity found in prostatic and seminal fluid. Uncritical use of the NADP+ assay might account for different seminal creatine kinase values reported in the literature. Interrelationships between ATPase, creatine kinase and zinc suggest that seminal ATPase is a prostatic secretory product while creatine kinase may be multiglandular in origin.     

The measurement of E and 19-hydroxy E prostaglandins in human seminal plasma

A method is described which measures the four main prostaglandins of human semen (PGE₁, E₂, 19-hydroxy PGE₁, and 19-hydroxy PGE₂). For routine measurements E₁ and E₂ are measured together as are 19-OH E₁ and 19-OH E₂. These are measured by forming oximes in aqueous solution. extraction, methylation and trimethyl silylation followed by gas chromatography. The method has sufficient sensitivity to measure the levels found in the majority of semen samples. The normal range in men with proven fertility was 90 to 260 μg/ml of 19-hydroxy Es and 30–200 μg/ml of Es.     

The measurement of E and 19-hydroxy E prostaglandins in human seminal plasma

A method is described which measures the four main prostaglandins of human semen (PGE₁, E₂, 19-hydroxy PGE₁, and 19-hydroxy PGE₂). For routine measurements E₁ and E₂ are measured together as are 19-OH E₁ and 19-OH E₂. These are measured by forming oximes in aqueous solution. extraction, methylation and trimethyl silylation followed by gas chromatography. The method has sufficient sensitivity to measure the levels found in the majority of semen samples. The normal range in men with proven fertility was 90 to 260 μg/ml of 19-hydroxy Es and 30–200 μg/ml of Es.     

Gas chromatographic determination of E prostaglandins in human amniotic fluid

A simple procedure is described for the extraction and purification of prostaglandins from amniotic fluid. Combined with gas chromatography with electron-capture detection, this provides a rapid and specific method for the determination of PGE in human amniotic fluid.     

The measurement of E and 19-hydroxy E prostaglandins in human seminal plasma.

A method is described which measures the four main prostaglandins of human semen (PGE1, E2, 19-hydroxy PGE1, and 19-hydroxy PGE2). For routine measurements E1 and E2 are measured together as are 19-OH E1 and 19-OH E2. These are measured by forming oximes in aqueous solution extraction, methylation and trimethyl silylation followed by gas chromatography. The method has sufficient sensitivity to measure the levels found in the majority of semen samples. The normal range in men with proven fertility was 90 to 260 mug/ml of 19-hydroxy Es and 30-200 mug/ml of Es.     

Isolation of leukotriene C4 from human seminal fluid

LTC4 was isolated and characterized from seminal fluid of seven human volunteers. A compound with a similar retention time to that of synthetic LTC4 was obtained using reverse-phase high performance liquid chromatography. The ultraviolet absorbance of the extracted substance was identical to synthetic LTC4. Furthermore this compound contracted the guinea pig ileum and lung parenchymal strip. Its effects were antagonized by the leukotriene antagonist FPL55712. It was concluded that LTC4 is present in human seminal fluid in very small amounts (about 100 ng/ejaculate). The possible physiological functions of LTC4 in the reproductive tract are discussed.     

Rapid and slow hydroxylators of seminal E prostaglandins

Human seminal fluid contains prostaglandin (PG) E1, PGE2, 19-hydroxy-PGE1 and 19-hydroxy-PGE2 in large and variable amounts. 19-Hydroxy-PGE1 and 19-hydroxy-PGE2 are formed from PGE1 and PGE2 by prostaglandin 19-hydroxylase, a cytochrome P-450 enzyme, in seminal vesicles. The hypothesis that genetic polymorphism of this enzyme might contribute to the variable concentrations of PGE1, PGE2, 19-hydroxy-PGE1 and 19-hydroxy-PGE2 was examined by analysis of seminal fluid of 40 normal men. E prostaglandins were measured with 17-phenyl-PGE2 as an internal standard by high-performance liquid chromatography on beta-cyclodextrin silica. Using the ratios of substrate/product, i.e., R1 = PGE1/19-hydroxy-PGE1 and R2 = PGE2/19-hydroxy-PGE2, as indicators of prostaglandin 19-hydroxylase capacity, a bimodal distribution of R values was found: nine men (23%) were slow hydroxylators (R1 greater than 0.45 and R2 greater than 0.45), while the remaining men were rapid hydroxylators (both R1 and R2 less than 0.45). Semen of slow hydroxylators and semen of the five most rapid hydroxylators (both R1 and R2 less than 0.10) differed in absolute amounts of PGE1 and PGE2 but not in 19-hydroxy-PGE1 and 19-hydroxy-PGE2. 20-Hydroxy-PGE1 and 20-hydroxy-PGE2 are formed from PGE1 and PGE2 by cytochrome P-450 in the vesicular glands and the ampullae of deferent ducts of the ram. Seminal fluid of five rams was analyzed for PGE1, PGE2, 20-hydroxy-PGE1 and 20-hydroxy-PGE2, and a large variation in substrate/product ratios was found. Polymorphism of cytochrome P-450 might contribute to variations in seminal prostaglandins in man and in sheep.     

Detection of prostatic tissue and seminal plasma peptides reacting with human seminal fluid acid phosphatase antibodies

IgG1 monoclonal antibody to purified seminal fluid phosphatase was raised by fusion of spleen cells from immunized mice with cell line Sp2/O-Ag 14 using simple method of screening for antiphosphatase antibody secreting clones. All molecular forms of catalytically active seminal fluid phosphatase and prostatic tissue phosphatase, resolved by chromatofocusing in pH gradient, react with this monoclonal antibody and with rabbit antiserum to purified seminal fluid phosphatase. Peptides of Mr 25,000 to 76,000 and of Mr 13,000 to 76,000 were adsorbed from the prostatic tissue extract and from seminal plasma on the monoclonal antibody-Sepharose column.     

Immunoreactive vasopressin in bull seminal fluid

Immunoreactive arginine vasopressin (irAVP) was measured in seminal fluid with and without extraction using a specific radioimmunoassay (RIA). A large fraction of irAVP was removed after extraction on octadecasilylsilica cartridges. The measured amount of irAVP corresponded to the levels found in blood plasma. Dilutions of seminal plasma extracts were parallel with the RIA standard curve. On reversed phase HPLC the extracted material coeluted with synthetic AVP. These findings suggest an identity of this immunoreactive material with intact AVP. During incubations of synthetic AVP and its analogue 8-D-arginine vasopressing (8-DAVP) in seminal plasma, immunoreactivity decreased considerably with the former peptide, while the concentration of 8-DAVP was not significantly altered.