Testing of Performance Enhancing Drugs James Rosenberger September 22, 2014.

Testing of Performance

Enhancing DrugsJames RosenbergerSeptember 22, 2014

Commonalities

Wes Welker - AdderallManny Ramirez – human chorionic gonadotropin

Lance Armstrong – human growth hormone, steroidsAlex Rodriguez – PrimobolanRyan Braun – Testosterone

http://nfl.blog.chatsports.com/files/2014/05/welker.jpg

http://elflowlatino.com/wp-content/uploads/2014/05/rex.jpg

http://www.qatar-tribune.com/data/20140720/gallery/27_2_1.jpg

http://www.forbeschina.com/upload/av6P0igR8S.jpghttp://rlindz35.files.wordpress.com/2012/08/braun-usatoday-net.jpeg

Ryan Braun

http://i.ytimg.com/vi/8UOcHEQ3z0U/0.jpg

http://i.ytimg.com/vi/8UOcHEQ3z0U/0.jpghttp://i.usatoday.net/sports/_photos/2011/09/25/MLB-games-matter-after-playoff-spots-clinched-A6DTHLR-x-large.jpg

Frowned Upon

• Uneven playing field• “How [am I] supposed to compete when others were

taking PEDS?” –Jeff Cirillo, 2002

• Health Side Effects• Acne• Impaired liver function• Increase in estrogen• Male pattern baldness

• Sets poor example for children• Shortcuts• Young people (Under 21) should NOT take steroids.

Doping

• Steroids/HGH/Blood Doping/Stimulants• What do they do?• Make muscles grow faster• Harmful side affects to health• Reduce fatigue• Recover from injuries quicker• Most major sports have banned them

• But how does it really work?• How are we stopping them?

http://upload.wikimedia.org/wikipedia/en/8/8f/WADA_logo.jpg

Steroids in Baseball

• Thomas Boswell, 1988• “The most conspicuous example of a

player [Jose Canseco] who has made himself great with steroids.” –CBS Late Night

• Truth without portfolio• Couldn’t publish this in a newspaper

• Beat Writers heard rumors, but could never whistle blow• Bob Nightengale, July 1995

• “Steroids Become an Issue…”Jose Canseco

http://rosachiropracticalexandria.com/clients/3050/images/Jose_Canseco_Chiropractor_Alexandria_VA.jpg

Great Home Run Race of 1998

Barry Bonds Record 2001 Season

http://en.wikipedia.org/wiki/Barry_Bonds#mediaviewer/File:Barry_Bonds_756_Ball.png

http://espn.go.com/media/classic/2001/0910/photo/sosamcgwire_ct.jpg

Steroids

• Definition: Refers to a class of hormones with a classic poly-phenol ring structure.• Catabolic Steroids (glucocorticoids)• Produced in the adrenal cortex or made

synthetically. Various metabolic function use to treat inflammation.

• Anabolic Steroids• Synthetic steroid hormone that

resembles testosterone in promoting the growth of muscle.

Gonanehttp://commons.wikimedia.org/wiki/File:Gonane.svg

Anabolic Steroids

• “Contain a synthetically manufactured form of the hormone testosterone”• Primary male sex hormone• An enhanced rate of protein

synthesis leading to muscle accumulation• Reason why males carry more muscle

mass than women• 11mg per day vs ¼ mg per day.

Testosterone

http://upload.wikimedia.org/wikipedia/commons/c/ce/Testosteron.svg

Gonane

http://commons.wikimedia.org/wiki/File:Gonane.svg

(Simple) Biochemistry of Anabolic Steroids

• Testosterone binds to intracellular receptor site in cytosol• Form Receptor complex and it

migrates to cell’s nucleus• Attaches to cell’s DNA• Activates transcription of

specific genes• Increase synthesis of two

primary contractile proteins, actin and myosin

ANABOLICS, 10th edition, William Llewellyn

Human Growth Hormones (HGH)

• Naturally occurring protein hormone• Excessive amounts give user

benefits by stimulating• Protein synthesis for increased

muscle mass• Bone growth for stronger bones• Breakdown of fat cells

• Use of HGH has become increasingly popular due to the difficulty of detection

http://www.nutrasanus.com/images/GrowthHormoneLevels.gif

Human Growth Hormones (HGH)

• Important Side-Effects• Stress on liver and

kidney• Swelling in the limbs• Migraines and

Headaches• High blood pressure• Bloating• Arthritis-like symptons

• But what makes testing so difficult?

http://www.nutrasanus.com/images/GrowthHormoneLevels.gif

http://images.publicradio.org/content/2013/01/01/20130101_hgh2_33.jpg

Human Growth Hormone Testing

Isoform Testing• At present, only blood tests have

found positive results• Natural HGH weighs 20 kDaltons• Synthetic HGH weighs 22

kDaltons• NFL & MLB: positive cutoff is 1.8• Downside:

• Short detection window (18-24 hours)

• 10,000 tests have been done since 2004 with only 14 positives.

HGH Testing

• A new method with LC-MS/MS is being tested• Measures IGF-1, Insulin-like

Growth Factor – 1• Blood test• IGF-1 is the response from an

active pituitary gland and a byproduct of HGH.

http://www.steroidtimes.com/wp-content/uploads/2010/08/nrd2359-f2-300x270.jpg

Liquid Chromatography

• Separation!• Stationary phase – Packed in column• Mobile phase – Solvent (based on polarity)

http://www.sec.psu.ac.th/web-board/content/view_img.php?id=667

Mass Spectrometry

• LC Product fed in• Ionized• Electric Field is altered

• Light particles get curved too much

• Heavy particles ignore field

• Tolerances are hard to find as detectors measure m/z• Retention Time and m/z ratio

can narrow down search considerably

http://www.chemguide.co.uk/analysis/masspec/masspec.GIF

T/E Ratio

• Testosterone/Epitestosterone• Epitestosterone is a byproduct

of testosterone• Normal ratio is 1:1• Positive test for NFL/MLB is 4:1• Carbon Isotope test follow up.• Pharmaceutical testosterone has

slightly more Carbon 12.

http://www.ftmguide.org/images/Tmolecule.gif

Concerns for Future

• Testing is expensive• Can we lower the price?• Use chemical reactions on

manufactured kits?• Avoid high priced machines

• Drugs are developing every year and finding a test will always lag behind• Lower tolerances, but avoid

false positives

http://quclms.files.wordpress.com/2013/06/dsc_0673.jpg

Real Life Examples

• Last week: NFL added HGH Testing into Drug Testing Policy• Possibly lower tolerances

• Olympians in Sochi• A passport of every drug test

they have had.• Gives testing facility a

comparison

• Drug Screening for Jobs

http://dmnewsi.files.wordpress.com/2013/11/drug-testing.jpg?w=1000

Questions?

http://dmnewsi.files.wordpress.com/2013/11/drug-testing.jpg?w=1000