http://pubs.acs.org/isubscribe/journals/cen/81/i46/html/8146sci2.html

As the UCLA researchers found out, THG is unstable under the conditions of the routine assay. "That's why we have been blind to it in previous testing," Catlin says.

When a new steroid such as THG is in a urine sample, it's not readily detectable using the standard algorithm, Catlin notes. However, there will be a telltale prominent peak in a region of the chromatogram where steroids generally appear.
Switching to an LC-MS/MS system made the process less dependent on the derivatization, Catlin notes. "We were actually able to get the retention time and mass spectrum of underivatized THG," he says. "It was difficult, but we got it."
That process turned out to be a good screen. It also gave the researchers a leg up once they found a suitable way to derivatize the material from the syringe, allowing them to determine the molecular ion of the steroid and begin to piece together what the molecule might be.
THG is thought to be derived from gestrinone, a synthetic steroid used to treat gynecological disorders. The two steroids differ by only four hydrogen atoms: THG has an ethyl group at C-17, whereas gestrinone has an ethynyl group there. THG also is structurally similar to trenbolone, a synthetic steroid that is used by bodybuilders and by U.S. ranchers in combination with estradiol to bulk up cattle.
While the UCLA chemists were still at the "pencil and paper stage," they were able to synthesize THG by simple hydrogenation of gestrinone, Catlin says. "We were able to show a match between the spectra of the synthesized THG and the THG from the syringe."
The synthesis set off a series of other investigations, he adds. As a confirmation, USADA had another testing lab synthesize THG. Catlin's lab also got help from Jane M. Strouse of the UCLA chemistry and biochemistry department's nuclear magnetic resonance imaging facility to characterize THG by NMR spectrometry. USADA further arranged to have an independent NMR expert do the same. USADA also arranged for THG to be given to a baboon. Urine samples collected from the baboon over a few days provided a close approximation of human urine to test the detection procedure and check for characteristic THG metabolites.

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DESIGNER STEROID ROCKS SPORTS WORLD
UCLA chemists identify steroid purportedly designed to avoid detection in routine tests
STEPHEN K. RITTER, C&EN WASHINGTON
Before mid-October, few people had heard of the synthetic steroid tetrahydrogestrinone (THG). That changed with the official announcement of the identification of THG and the development of a test method to screen urine samples for it, which altogether is being hailed by sports-governing authorities as a milestone in the 40-year history of international antidoping efforts. Meanwhile, a federal investigation is ongoing to determine if there was a conspiracy to synthesize and distribute THG or other unapproved or controlled substances.

STEROID PRIMER Tetrahydrogestrinone is a new synthetic steroid similar to the synthetic steroids gestrinone and trenbolone. Testosterone, the main male sex hormone, and its precursor, androstenedione, are natural compounds in the cascade of steroids derived from cholesterol.	
	
THE MYSTERIOUS SYRINGE is the first time in Catlin's 20-year career as director of the UCLA lab that a drug sample has turned up out of the blue. He suspects that a few underground labs are always attempting to design steroids and other drugs that can't be detected. "Our lab has been interested in designer steroids for some time, and we know the theory behind how one can make them to be undetectable. We have been doing some research trying to find them, but the vast number of possibilities you are looking for makes this a difficult project."
About a year ago, Catlin's group did find a previously undetected steroid in a single urine sample from a cyclist, he notes. The steroid, norbolethone, is not exactly a designer steroid like THG. Rather, it was developed by a drug company 40 years ago, but it never made it to market. "Someone out there was reading the old literature, which basically says norbolethone is very potent," Catlin says. "They must have made it. Then we found it in a urine sample." Norbolethone was added to the IOC/WADA list of prohibited substances and methods earlier this year.
Catlin believes it's likely that THG was designed specifically to avoid detection, but he acknowledges that it could just be a coincidence that the steroid's chemical properties made it undetectable. His group has gone back over spectra of previous samples looking for signs of THG, but they have not seen it, suggesting that the steroid has been used for only a short period of time.
"The underlying issue of the THG story is that we have found a steroid that heretofore we did not know about," Catlin says. "That points to some sort of industry--of one or more people--who are involved in developing, synthesizing, and distributing steroids. Getting on top of that is something that sports authorities are going to have to deal with proactively rather than after the fact."
The THG saga suggests that doping in sports will continue to be a cat-and-mouse game. As scientists attempt to develop better testing procedures, unscrupulous athletes will seek out other ways to cheat and, if caught, will continue to insist that their infractions don't violate the "spirit of the rules." Sports authorities probably will continue to be slow-footed in reacting because of the billions of dollars of revenues generated worldwide by professional and Olympic sports. Heading into the 2004 Summer Olympic Games next year in Athens, Greece, those involved on all sides of the doping issue are going to remain on edge.