Steroids and their diverse effects Nathan Kenyon Institute for Chemistry Literacy through...

Steroids and Steroids and their diverse their diverse

effectseffectsNathan KenyonNathan Kenyon

Institute for Chemistry Literacy through Institute for Chemistry Literacy through Computational ScienceComputational Science

University of Illinois College of Medicine at University of Illinois College of Medicine at Urbana-ChampaignUrbana-Champaign

©2011 University of Illinois Board of Trustees •

http://islcs.ncsa.illinois.edu/copyright

About this unitAbout this unit Author of original unitAuthor of original unit: Nathan Kenyon: Nathan Kenyon

Grade LevelGrade Level: High school chemistry, biology teachers: High school chemistry, biology teachers

Rationale of the Unit Rationale of the Unit The unit gives background information on steroid hormones, The unit gives background information on steroid hormones, explores their importance in human physiology, and looks at explores their importance in human physiology, and looks at the dangers of using synthetic anabolic steroids. The lesson the dangers of using synthetic anabolic steroids. The lesson also includes instructions for creating a testosterone molecule also includes instructions for creating a testosterone molecule using ChemSketch and then calculating its UV-absorption using ChemSketch and then calculating its UV-absorption spectra using WebMO.spectra using WebMO.

Use(s) of the UnitUse(s) of the Unit Intended for high school chemistry or biology teachers, who Intended for high school chemistry or biology teachers, who wish to use computational tools in the classroom. Also suitable wish to use computational tools in the classroom. Also suitable for introductory college biology. It could also be used directly for introductory college biology. It could also be used directly by students for self-study.by students for self-study.

Content of the Unit Content of the Unit Background information on steroid molecules, exercise in Background information on steroid molecules, exercise in creating biological molecules, calculating their propertiescreating biological molecules, calculating their properties©2011 University of Illinois Board of

Trustees • http://islcs.ncsa.illinois.edu/copyright

One reason for using One reason for using steroids:steroids:



The effects do not last The effects do not last forever….forever….



A reason to avoid steroid A reason to avoid steroid use:use:

Mark McGwire testifying in a 2005 congressional hearing on his alleged steroid use (Getty Images, 2005)



Major League Baseball on Major League Baseball on performance enhancing performance enhancing

drugsdrugs[Commissioner Bud Selig] “a reminder to [Commissioner Bud Selig] “a reminder to

everyone in baseball -- under our current drug everyone in baseball -- under our current drug program, if you are caught using steroids program, if you are caught using steroids and/or amphetamines, you will be punished. and/or amphetamines, you will be punished. Since 2005, every player who has tested Since 2005, every player who has tested positive for steroids has been positive for steroids has been suspendedsuspended for for as much as 50 games. Eradicating as much as 50 games. Eradicating performance-enhancing substances from the performance-enhancing substances from the game of baseball has been my first priority game of baseball has been my first priority over the past decade.”over the past decade.”

(MLB.com, 2009)(MLB.com, 2009)©2011 University of Illinois Board of


Reasons for using Reasons for using performance-enhancing performance-enhancing drugs:drugs:Build mass and strength of Build mass and strength of muscles and/or bonesmuscles and/or bones

Increase delivery of oxygen to Increase delivery of oxygen to exercising tissuesexercising tissues

Decrease painDecrease pain

Reduce weightReduce weight

Hide use of other drugsHide use of other drugs



promote growthpromote growth promote sex gland developmentpromote sex gland development affect salt/water balanceaffect salt/water balance suppress the immune systemsuppress the immune system reduce inflammationreduce inflammation involved in reproductive cyclesinvolved in reproductive cycles

Steroids can have a variety of functions in various body tissues:



Steroid SynthesisSteroid Synthesis consist of a fused four-ring structureconsist of a fused four-ring structure fat-soluble fat-soluble steroids are synthesized in the steroids are synthesized in the adrenal adrenal

glands glands or or gonadsgonads hormones travel in bloodstream to target hormones travel in bloodstream to target

tissuestissues



Steroid Ring StructureSteroid Ring Structure very common in biological very common in biological

compoundscompounds ““rings of life”rings of life” three cyclohexanes (six carbons) and three cyclohexanes (six carbons) and

one cyclopentane (five carbons)one cyclopentane (five carbons) seen in vitamins, drugs, hormones seen in vitamins, drugs, hormones



A brief exerciseA brief exercise Use Use WebMOWebMO to calculate quantities associated to calculate quantities associated

with molecular orbitals of cyclohexane and with molecular orbitals of cyclohexane and benzenebenzene

1.1. Open up Chemsketch in your browserOpen up Chemsketch in your browser

2.2. On the right side click on “cyclohexane” to On the right side click on “cyclohexane” to select a molecule to paste and then left click in select a molecule to paste and then left click in the drawing areathe drawing area



1.1. Under file menu click “save as” and save as a .mol Under file menu click “save as” and save as a .mol file using “save as type”file using “save as type”

2.2. Use cyclohexane as name of fileUse cyclohexane as name of file

3.3. Login in to WebMO using the username and Login in to WebMO using the username and password providedpassword provided

4.4. Click on “import molecule” and hit browse to find Click on “import molecule” and hit browse to find cyclohexane.molcyclohexane.mol

5.5. Click on open Click on open

-a 3D cyclohexane should appear in the -a 3D cyclohexane should appear in the screenscreen



Click the blue “continue” arrow in the lower right side of the Build Molecule window.

Choose Gaussian as the computational engine. Choose “first available” in the select server pulldown Menu, click the blue “continue” arrow. Type in/Choose the following:

Job Name: CH4PM3Calculation: Geometry OptimizationTheory: PM3Basis Set: Basic: 321G (or accept default)Charge: 0Multiplicity: Singlet

Click on the blue “continue” arrow. You should now see your job listed.

Click Refresh (every ~5 seconds) until you see that your job is Complete (under Status).

Click on the hyperlinked name (CH4PM3) to open the “View Job” window.

Choose the Select arrow (4th icon down on left)

When the job is done, click on the filename and use the rotate tool to investigate the structure.



Click on one of the C atoms in the structure (the other atoms and bonds will “fade”)

Shift and Click the C atom (both atoms are now highlighted)

Discern bond lengths, hybridizaion, charge, etc.

Bond length: 1.314 A Hybridization: Sp3 Charge: 0

CyclohexaneCyclohexane



BenzeneBenzene Click on one of the C atoms in the structure

(the other atoms and bonds will “fade”) Shift and Click the C atom (both atoms are

now highlighted) Discern bond lengths, hybridizaion, charge,

etc.

Bond length: 1.314 A Hybridization: Sp3 Charge: 0



Benzene vs. Cyclohexane partial Benzene vs. Cyclohexane partial chargescharges

carbon carbon numbenumbe

rr

benzenebenzene cyclohexcyclohexaneane

11 --0.000060.00006

2 2

--0.0000570.000057

22 --0.000060.00006

33

--0.000058 0.000058

33 --0.000000.00000

88

--0.0000350.000035

44 0.000070.0000711

0.0000920.000092

55 0.000070.0000711

0.0000920.000092

66 --0.000000.00000

88

--0.0000350.000035

Dipole Dipole momemome

ntnt

0.0016 0.0016

DebyeDebye0.00160.0016

DebyeDebye

cyclohexane

benzene

Partial Charges



Questions (Benzene vs. Questions (Benzene vs. Cyclohexane)Cyclohexane)

Based on the data collected from Based on the data collected from WebMO, what are some differences WebMO, what are some differences between benzene and cyclohexane? between benzene and cyclohexane? similarities?similarities?



Steroid precursor: Steroid precursor: cholesterolcholesterol



What is cholesterol?What is cholesterol? precursor to steroid compoundsprecursor to steroid compounds a real biological molecule, not simply a fatty a real biological molecule, not simply a fatty

buildup of plaquebuildup of plaque lipid-like, found in cell-membranes, necessary for lipid-like, found in cell-membranes, necessary for

membrane integritymembrane integrity high levels in blood plasma associated with high levels in blood plasma associated with

atherosclerosis (deposits in vessels can lead to atherosclerosis (deposits in vessels can lead to disease)disease)



Cholesterol conversion Cholesterol conversion



TestosteroneTestosterone Principal male sex hormone and Principal male sex hormone and

anabolic steroidanabolic steroid



Androstenedione Androstenedione

Nutritional supplement used by athletesNutritional supplement used by athletes Precursor to testosteronePrecursor to testosterone



Conversion of Androstenedione to Conversion of Androstenedione to TestosteroneTestosterone

17-Beta-hydroxysteroid dehydrogenase

-Thus, an athlete may take androstenedione to increase testosterone levels in the body



Examination of Examination of Androstenedione vs. Androstenedione vs.

TestosteroneTestosterone

AndrostenedioAndrostenedionene

TestosteronTestosteronee

**note the polarity change on the oxygen



17-beta-hydroxysteroid 17-beta-hydroxysteroid dehydrogenase deficiencydehydrogenase deficiency

Produce male pseudohermaphroditismProduce male pseudohermaphroditism-ambigious genitalia-ambigious genitalia

Enzyme deficiency leads to low levels of Enzyme deficiency leads to low levels of testosteronetestosterone



Dangers of SteroidsDangers of Steroids



Dangers of Steroid UseDangers of Steroid Use AcneAcne

Liver abnormalitiesLiver abnormalities

Testicular atrophy ("Honey, I shrunk the testicles"), Testicular atrophy ("Honey, I shrunk the testicles"), subsequent infertilitysubsequent infertility

Masculinizing effects in womenMasculinizing effects in women

Adolescents: Premature closure of the growth plates Adolescents: Premature closure of the growth plates in bone resulting in shorter adult staturein bone resulting in shorter adult stature

Breast enlargement in menBreast enlargement in men

Mood changes (“Roid Rage”)Mood changes (“Roid Rage”)



Another important steroid: Another important steroid: AldosteroneAldosterone

Important for water retention

Acts on kidney to increase sodium absorption (water follows)



Contrast the two hormonesContrast the two hormones

Aldosterone: water retention

Testosterone: development of secondary sex characteristics

Changing the sidechains and functional groups can lead to vastly different effects in the body!



Aldosterone MechanismAldosterone Mechanism

Hormone binds to Hormone binds to collecting collecting duct/distal tubule duct/distal tubule cells in kidney cells in kidney nephronnephron

Principal cells Principal cells absorb more Naabsorb more Na++, , water and secrete water and secrete KK++

Increases blood Increases blood volume and hence volume and hence blood pressureblood pressure

Renin-Angiotensin Renin-Angiotensin SystemSystem



Why does water follow?Why does water follow?

More NaMore Na++ is being reabsorbed into is being reabsorbed into collecting duct due to aldosteronecollecting duct due to aldosterone

NaNa+ + is positive and water is partially is positive and water is partially negative due to oxygennegative due to oxygen

NaNa+ +



CortisolCortisol Corticosteroid released in response to Corticosteroid released in response to

stressstress Increases blood pressure and blood Increases blood pressure and blood

sugar, reduces immune responsessugar, reduces immune responses



CortisolCortisol Elevated levels seen during stressful Elevated levels seen during stressful

situations, dieting, sleep deprivationsituations, dieting, sleep deprivation High levels of cortisol bad for healthHigh levels of cortisol bad for health Levels fluctuate throughout dayLevels fluctuate throughout day

Cortisol Levels Throughout the Day

024

681012

141618

4AM 6AM 8AM 10AM Noon 2PM 4PM 6PM 8PM 10PM Mid 2AM

Stress

Normal ©2011 University of Illinois Board of Trustees •


Other steroid compoundsOther steroid compounds

gonane estrane

pregnane

All available in chemsketch under “templates”



Testing for steroid useTesting for steroid use Measure testosterone in urine using Measure testosterone in urine using gas gas

chromatography/mass spectrometrychromatography/mass spectrometry

Endogenous testosterone and injected Endogenous testosterone and injected testosterone differ in their absorption of testosterone differ in their absorption of lightlight

Calculating Calculating UV-absorption spectraUV-absorption spectra allows allows scientists to determine whether an athlete scientists to determine whether an athlete has banned substances in the bodyhas banned substances in the body

National Collegiate Athletic Association National Collegiate Athletic Association World Anti-Doping AgencyWorld Anti-Doping Agency ©2011 University of Illinois Board of


Steroid DetectionSteroid Detection



Let’s build a steroid!Let’s build a steroid! Open up chemsketch in your browserOpen up chemsketch in your browser On the right side, click on the six-carbon On the right side, click on the six-carbon

ring structure (cyclohexane)ring structure (cyclohexane)



Building a Steroid…..Building a Steroid….. Move the cursor to the center of the Move the cursor to the center of the

screen and click to draw one hexane screen and click to draw one hexane moleculemolecule

Move the cursor to the left or right and Move the cursor to the left or right and click to add another hexaneclick to add another hexane



Building a Steroid…..Building a Steroid….. Click on the pentane button on the Click on the pentane button on the

right side to finish the fused carbon right side to finish the fused carbon ring structurering structure

On the left click the “O” for oxygen On the left click the “O” for oxygen and then click the “draw chains” at and then click the “draw chains” at the top toolbarthe top toolbar



Click on the left bottom-most carbon to Click on the left bottom-most carbon to add the “O” to the hexaneadd the “O” to the hexane

Click the bond to oxidize itClick the bond to oxidize it Continue using this function to add the Continue using this function to add the

necessary functional groups to create necessary functional groups to create testosterone (note: you must be in “add testosterone (note: you must be in “add chains” mode)chains” mode)

Import into WebMO and make Import into WebMO and make calculationscalculations

Finished product shown below:Finished product shown below:



Calculating absorbance spectra of Calculating absorbance spectra of testosteronetestosterone

Access the following Web site: Access the following Web site: http://neo.beckman.uiuc.edu

Log in as “Guest.” Password: Guest.Log in as “Guest.” Password: Guest. Click on the main server.Click on the main server. Click on NEW JOB. When the pop-up work space Click on NEW JOB. When the pop-up work space

appears, click on import molecule. appears, click on import molecule. Click on BROWSE.Click on BROWSE. Click on the ChemSketch file that contains the Click on the ChemSketch file that contains the

correct structure for testosteronecorrect structure for testosterone

Click IMPORT MOLECULE. Click IMPORT MOLECULE.

Click on OPEN EDITOR.Click on OPEN EDITOR.

Click on CLEAN-UP>ADD HYDROGENS.Click on CLEAN-UP>ADD HYDROGENS.©2011 University of Illinois Board of



Close editor and click on the blue CONTINUE arrow.Close editor and click on the blue CONTINUE arrow.

Select MOPAC and click on the blue arrow again.Select MOPAC and click on the blue arrow again.

Type in the following:Type in the following: Job Name: C16H22O3 (true)Job Name: C16H22O3 (true)

Calculation: Geometry OptimizationCalculation: Geometry OptimizationTheory: PM3Theory: PM3Charge: 0Charge: 0Multiplicity: SingletMultiplicity: Singlet

Submit the job by clicking on the blue continue arrow.Submit the job by clicking on the blue continue arrow.

When the job is done, click on the filename and use When the job is done, click on the filename and use the rotate tool to investigate the structure. the rotate tool to investigate the structure.




Click on NEW JOB USING THIS GEOMETRY. Click Click on NEW JOB USING THIS GEOMETRY. Click the blue CONTINUE arrow and select GAUSSIAN as the blue CONTINUE arrow and select GAUSSIAN as the computational engine. Click the arrow again and the computational engine. Click the arrow again and select/type in:select/type in:

Job name: C16H22O3Job name: C16H22O3Calculation: UV-Vis spectrumCalculation: UV-Vis spectrumTheory: Hartree-FockBasis Set: Other (Cancel the pop-up window)Charge: 0Multiplicity: Singlet

Click on the ADVANCED tab. Type in ZINDO in the ADDITIONAL KEYWORDS window.

Click on the PREVIEW tab; then click GENERATE. Edit the first line to read:

#N CIS=(NStates=10)/ZINDO(i.e., remove “null” and the space after the slash)



Submit job by clicking on SUBMIT JOB on the left side. Click OK in the pop-up window.

When the job is complete, click on the filename and scroll down to view the EXCITED STATES table. Near the bottom of the table, click on the magnifying glass to view the calculated spectrum. In the SPECTRUM VIEWER window, use our mouse and place the cursor on the top of the peak and record the wavelength at maximum absorbance (lmax = displayed in the upper right of the SPECTRUM VIEWER window):



Questions for reviewQuestions for review Where in the body are steroids produced? Where in the body are steroids produced?

What is the purpose of taking a synthetic Androstenedione?What is the purpose of taking a synthetic Androstenedione?

What are some of the negative effects of taking anabolic What are some of the negative effects of taking anabolic steroids?steroids?

What information can be taken from the absorbance spectra of What information can be taken from the absorbance spectra of testosterone?testosterone?



Questions for reviewQuestions for review

Use WebMO or ChemSketch to draw Use WebMO or ChemSketch to draw Cortisol (shown below), a hormone that Cortisol (shown below), a hormone that increases blood sugar and blood glucose in increases blood sugar and blood glucose in reponse to stress.reponse to stress.

Why would prolonged high levels of cortisol Why would prolonged high levels of cortisol be harmful?be harmful? ©2011 University of Illinois Board of


Answers to QuestionsAnswers to Questions Where in the body are steroids produced? Where in the body are steroids produced?

adrenal glands, gonadsadrenal glands, gonads

What is the purpose of taking a synthetic Androstenedione?What is the purpose of taking a synthetic Androstenedione?increase testosterone levels (increase muscle mass)increase testosterone levels (increase muscle mass)

What are some of the negative effects of taking anabolic What are some of the negative effects of taking anabolic steroids? steroids?

acne, testicular atrophy, mood swings, breast acne, testicular atrophy, mood swings, breast enlargement enlargement (men), (men), masculinization in womenmasculinization in women

Why are prolonged levels of cortisol harmful?Why are prolonged levels of cortisol harmful?High levels of cortisol cause fat stores and excess High levels of cortisol cause fat stores and excess

circulating fat to be relocated and deposited deep in the circulating fat to be relocated and deposited deep in the abdomen, which left unchecked can develop into obesity. abdomen, which left unchecked can develop into obesity.

Hypertension (high blood pressure), Hyperlipidemia Hypertension (high blood pressure), Hyperlipidemia (elevated lipids), and Hyperglycemia (elevated (elevated lipids), and Hyperglycemia (elevated

glucose) glucose) have been linked to have been linked to elevatedelevated cortisol levels cortisol levels



ReferencesReferences

http://www.unm.edu/~lkravitz/Article%20folder/stresscortisol.html

http://www.pubmedcentral.nih.gov/http://www.pubmedcentral.nih.gov/articlerender.fcgi?artid=1402217articlerender.fcgi?artid=1402217

http://www.howstuffworks.com/athletic-drug-test.htm



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