Anabolic activity assay

In addition to the classical tests usually applied for the assay of androgenic and anabolic activities, another method is reported by the same workers for the evaluation of androgenic properties of progestational compounds in the rat by the female fetal masculinization test [201]. It should be pointed out that inversion of configuration at C-10 does not necessarily bring about abolition of androgenic activity, in view of the new theory, as will be seen later. 

A -Testosterone (S-63) and A -Sa-dibydrotestosterone (D-48) show increased androgenic and anabolic activities in the rat when administered orally and decreased activities when administered subcutaneously. Chick comb assays for androgenic activity show that the A -compounds are more active than their parent compounds. However, in the mouse androgen assay A -5a-dihydrotestosterone is more active and A -testosterone is less active than their parent compounds. 

One of the best efforts to compare the anabolic activity of many steroids which have been discussed is that of Albanese and coworkers (1), who devised a careful means of comparing a wide series of steroids. They employed nitrogen balance studies in appropriate patients. They then expressed the steroid protein activity index, the SPAI, on a comparative basis. The antianabolic or catabolic adrenal cortical steroids all have negative values by this assay. Table II shows comparative SPATs of seven of the steroids which have been discussed. It can be seen by this index that testosterone propionate is the least active and stanozolol the most active. Further reports from Albanese and coworkers will be anticipated with interest. 

It is noteworthy that for the Lesch-Nyhan syndrome fibroblasts, the assay in the presence of TTP furnished the bulk of the radioactivity in the nucleoside fraction rather than in the nucleotide fraction, in contrast to normal cell extracts in which the bulk of the radioactivity is found in the nucleotide fraction. Two possible explanations may be entertained to explain this observation. First, in the HGPRT-deficient system the nucleotidase activity is far in excess over the HGPRT activity, and presumably the residual nucleotidase activity, not inhibited by TTP, is sufficient to degrade the relatively small amount of nucleotide formed. Secondly, the possibility should be considered that the accumulation of radioactivity in the nucleoside fraction reflects the anabolic activity of nucleoside phospho-rylase reacting the purine base with ribose-l-phosphate to form the nucleotide by an alternative pathway. However, this latter explanation seems to be invalid in view of the absence of the suitable substrate, ribose-l-phosphate, from the incubation system, and by the linearity... 

This chapter describes the use of the HPLC method to assay the activity of several enzymes simultaneously. The examples include several different enzymes that can use the same substrate and form the same product, a single enzyme that can use different substrates to form different products and two different activities using the same substrate to form different products. In another example the use of the HPLC method to study metabolic pathways is described through a series of reconstitution studies, and finally the HPLC method is applied to the anabolism of adenosine. 

The 2 -azido group of cytarazid renders the nucleoside more resistant to deamination to the 2 -azidouridine derivative (153) by deoxycytidine deaminase, but was also observed to reduce substrate affinity for the deoxycytidine kinase necessary for anabolic phosphorylation to the active cytarazid 5 -triphosphate [180]. Conversely, cytarazid was a more potent inhibitor of the target DNA polymerases a and K = 0.6 and 0.7 //M, respectively) than the parent ara-C (.A = 10 and 17 fiM, respectively), and the dissimilar spectrum of antitumour activity exhibited by the two compounds was attributed to differences in stability, metabolic activation and inhibitory potency [180, 181]. Interestingly, the instability of cytarazid to thiols present in the assay media, was commented on but not pursued [180]. In view of previous discussions concerning the bioreduction of AZT... 

Discovery of NM-107 (6), the first HCV NS5B nucleoside polymerase inhibitor from Idenix Pharmaceuticals (now Merck), in combination with the discovery of 2 -deoxy-2 -fluorocytidine (7) by the Pharmasset research team led to the discovery of PSI-6130 (8). The metabolism study of PSI-6130 showed that PSI-6130 was anabolized to the active triphosphate, PSI-6130-TP (9), and two other metabolites, PSI-6206 (10, R02433, GS-331007) and PSI-6206-TP (11, PSI-7409, GS-461203). 2 The biological evaluation revealed that the triphosphate of PSI-6206 (11, PSI-7409, GS-461203) was a potent and selective inhibitor of HCV NS5B polymerase (IC90 = 0.52 pM), while PSI-6206 was inactive in the HCV rephcon assay (EC90 > 100 Subsequent... 

The development of analytical assays enabling the comprehensive detection of AAS in sports drug testing samples has required the consideration of the commonly rather extensive metabolism of these xeno-biotics. Principle metabolic reactions of synthetic anabolic steroids are closely related to those reported for testosterone (see section 6.1.2.2), with major pathways controlled by various enzymes such as members of the cytochrome P450 system, 17 3-hydroxysteroid dehydrogenase, 3a-/3P-hydroxysteroid dehydrogenase and 5a-reductases, which are primarily responsible for the conversion of testosterone into active... 

---