ready for take-off – crystallography in undergraduate education

Ready for Take-Off –

Crystallography in Undergraduate Education

August 29, 2012 1

Ready for Take-Off – Crystallography in Undergraduate Education

August 29, 2012 2

Dr. Michael Ruf Product Manager – Crystallography Bruker AXS Inc. Madison, WI, USA Dr. Roger Sommer Assistant Professor Department of Chemistry DePaul University Chicago, IL, USA Dr. Nigam Rath Director of the X-ray Diffraction Lab Department of Chemistry & Biochemistry University of Missouri St. Louis, MO, USA

• Introduction - Michael Ruf

• Experiences at DePaul University, Chicago - Roger Sommer

- CHE480: Special Topics in Chemistry: X-ray Crystallography

- From growing and handling crystals to preparing reports of experimental data for publication

• Experiences at University of Missouri, St. Louis - Nigam Rath

- Organic and inorganic chemistry experiments: syntheses, crystallization and structure determination of organic and inorganic compounds

Ready for Take-Off – Crystallography in Undergraduate Education

August 29, 2012 3

Is there a steep learning curve in crystallography?

August 29, 2012 4

• Crystallization

• X-rays

• Reciprocal space

• Fast Fourier Indexing

• Data collection strategies

• Kappa geometry

• Patterson methods

• SIRAS, SAD, MAD …

August 29, 2012 5

• Should we require that students understand structure factors and comprehend the concept of reciprocal space to be able to determine a structure?

• How can we interest students in crystallographic methods for structure determination?

• Can we interest students in crystallography by providing easy access to structure determination?

• Can we make it easier?

• Is automation a viable option?

• Can automation provide reliable and accurate structural information?

How can we interest students in crystallography?

August 29, 2012 6

• It is possible to take good quality pictures without any knowledge of

- Exposure times

- F-stops

- CMOS or CCD technology

- Image processing

- Or even one look at the users manual

Point and shoot in photography

VS.

August 29, 2012 7

• Is it possible to successfully apply this concept to structure determination?

• Would this concept lower the barrier to getting students interested in crystallography?

Point and shoot in crystallography?

VS.

August 29, 2012 8

SMART X2S benchtop diffractometer

The SMART X2S is

• a true benchtop system which can be placed anywhere in your laboratory

• extremely compact, lightweight and completely air-cooled – no cooling water is needed

• Requires single phase power only and draws ten times less power than a conventional diffractometer

August 29, 2012 9

SMART X2S benchtop diffractometer

• Completely automated operation

- Sample mounting and alignment

- Data collection

- Structure solution

- Report generation

- High success rate

• 100% data compatibility with APEX2 – the crystallography software suite

SMART X2S in a teaching environment

Roger Sommer

DePaul University

Department of Chemistry

August 29, 2012 11

In your opinion, what is the major hurdle to teaching crystallography to undergraduates?

a) Growing crystals

b) Availability of instrumentation

c) Safety

d) Need for local expertise

e) Availability of time in curriculum

Audience Poll

CHE480: Special Topics in Chemistry X-ray Crystallography

• 2 credit hour (1/2 regular course) – 1.5 hours lecture once a week for 10 weeks – 6 Masters, 3 senior undergrads

• Specific objectives of the course is to teach students how to: – Grow and handle crystals. – Run an experiment using the Bruker SMART X2S – Refine model to best fit the data. Evaluate the quality of the

structure based on common metrics. – Make useful high quality graphics of solid state structures. – Prepare report of experimental data for possible

publication.

My Objectives

• Students become independent users of SMART X2S

– Able to complete a routine experiment without direct intervention of crystallographer (instructor)

• Teach chemistry through crystallography

– CHE321 substitute (Int. Inorg. Chem.)

– Chemical intuition guides model refinement

Course Schedule Date Topics Events

9/13/2010

A look at the instrument, setup of experiment, how

data gets collected, a basic look at Bragg’s law, start a

dataset, look at unit cell, Miller indices, diffraction

planes. Get started using software.

9/20/2010

Crystal systems, Bravais lattices, space groups,

symmetry operations in unit cell, systematic absences,

look at international tables, evaluating crystal quality

under microscope.

Project 1 selection

9/27/2010

Completing and refining structure model, look at

extended structure (intermolecular interactions),

microscope/lab time

10/4/2010 Growing crystals, project selection/start crystallizations Project 1 due,

Project 2 selection

10/11/2010

Structure refinement - atom identification

project time, discussion of examples/specific issues

with student projects

10/18/2010 Techniques for dealing with solvated structures,

disorder and other problems, project 2 selection. Project 3 selection

10/25/2010 Project time/discussions Project 2 due

11/1/2010 Project time/discussions

11/8/2010 Project time/discussions

11/15/2010 Project time/discussions

11/22/2010 Project 3 due

Project 1: Do a structure of a pre-made crystal

• In the lab – Assess sample quality and select viable

specimen (microscope/X2S) – Mount crystal and start data collection – Observe unit cell determination and check

CSD for matches

• Homework – Refine structure

• X2S, OLEX2, or APEX2/XShell with instructor help

– Practice generating useful graphics – Write useful captions

• Report – One paragraph description of structure – Quality graphic of asymm. unit or molecule – Packing diagram from Hg including

characteristic symm. element

nickel sulfate taurine hexamethylenetetramine sucrose copper(II)acetate dihydrate vanadyl sulfate

Project 1: Instructor input

• 30-45 minutes outside lecture per student

– Microscope training and crystal selection/mounting

• 30 minutes Lecture time introducing software

• 3 hours of office hours troubleshooting structure refinement

Student illustrations of symmetry and Miller planes from Mercury

Si on 2 fold axis in P21212 222 plane

Mercury CSD 2.0 - New Features for the Visualization and Investigation of Crystal Structures C. F. Macrae, I. J. Bruno, J. A. Chisholm, P. R. Edgington, P. McCabe, E. Pidcock, L. Rodriguez-Monge, R. Taylor, J. van de Streek and P. A. Wood, J. Appl. Cryst., 41, 466-470, 2008 http://www.ccdc.cam.ac.uk/

Project 1 pictures: Illustrate symmetry elements

Figure: The unit cell of NiSO4H2O. The cell has

the dimensions 6.78597Å x 6.78597Å x

18.2992Å and the green lines indicate the

location of the 2-fold rotation axes through the

sulfur and nickel atoms of the compound. Figure 1: 2-Fold symmetry with (0.5-x,

0.5+y, 0.5-z) transformation was used to

generate above figure.

J. Appl. Cryst., 41, 466-470, 2008

Project 2: Grow your own crystals, improve graphics/descriptions

• In the lab – Students grow, select, and mount

their own crystals

• Homework – Refine structure using OLEX2, ShelXL

– Add appropriate labeling scheme and improve graphic/caption quality

• Report – Similar to 1 but better quality

graphics, improved language/descriptions

1) Dolomanov, O.V.; Bourhis, L.J.; Gildea, R.J.; Howard, J.A.K.; Puschmann, H., OLEX2: A complete structure solution, refinement and analysis program J. Appl. Cryst. 2009, 42, 339-341

2) SHELXL, G.M. Sheldrick, Acta Cryst. 2008 A64, 112-122

Project 2: Instructor input

• Lectures and discussion on refining structure and improving reports

• ~30-45 min/student on crystallizations

- Students spent more time • Office hour questions on

software and refinement

• 5-10 minutes per student for access to X-ray lab

Project 3: Finishing the structure • In the lab (optional)

– Some students used pre-collected data – Some got extensions to grow crystals of new compounds over winter break

• Homework

– Refine structure to convergence (OLEX2) – Merge and write .cif file (OLEX2) – Edit and complete .cif file (enCIFer) – Use CheckCIF and address as many issues as possible

• Report

– Quality graphics and description, "completed" .cif file

– A copy of the "initial" and "final" CheckCIF report – Standard tables for a paper

CIF applications. XV. enCIFer: a program for viewing, editing and visualising CIFs. F. H. Allen, O. Johnson, G. P. Shields, B. R. Smith, M. Towler, J. Applied Cryst., 2004 , 37, 335-338.

Student response

• Enthusiastic and engaged

• “Hands-on”

• “My own Data”

• “Practical application”

• Students used compounds from their own Master’s project

• Several students ready to publish

The numbers from CHE480

• Samples studied: ~50

– Includes many samples from student research that did not diffract

• Datasets collected: ~25

• Structures: 20

• New structures: 10

Students wanted more of…

• Discussion of Cambridge Structural Database

• A CSD-based project

– Analyze intermolecular interactions etc.

Student frustrations

• Growing crystals

• “A lot of software to learn…”

Student engagement

Kristy On Tue, Nov 30, 2010 at 6:14 PM, Kristina Streu < wrote: Dr. Sommer, I'm in the lab playing around with some of the crystals of the 2-acetamidobiphenyl that we grew. I got one loaded onto a mount and I started the experiment. However the instrument has stopped while loading the sample. It has been showing the "please insert the sample" screen for about 5 minutes, and has reported a loading error. I was hoping you would have some insight as to what I might need to do to retrieve the sample and fix this loading issue. In the loading process the instrument seems to have stalled at teh point where the sample has been taken down, however the door has not closed. My number is if you wanted to call me, I will be in the lab for a while working on Project 3 and some research. Or you can e-mail me back, whatever is convenient for you. Thanks, Kristy

Student engagement

Pca21

From: Kristina Streu [mailto: ] Sent: Tuesday, November 30, 2010 6:42 PM To: rsommer@depaul.edu Subject: Re: Crystals Dr. Sommer, Tom came in and advised me to try turning the instrument on and letting it re-initialize. This fixed the problem. The 2-acetamidobiphenyl crystal is running just fine now. Hopefully we get something from it. Kristy

Summer research with the X2S

• Two students in Sommer Lab

– Rising sophomores

• Ready to “go solo” within a week

• Three more structures for colleagues

Summer research results presented as a poster Spring ACS meeting 2011

August 29, 2012 30

Poll Results

In your opinion, what is the major hurdle to teaching crystallography to undergraduates?

a) Growing crystals

b) Availability of instrumentation

c) Safety

d) Need for local expertise

e) Availability of time in curriculum

Crystallography Experiments for

Undergraduate Laboratory

Nigam P. Rath,

Department of Chemistry and Biochemistry and Center for Nanoscience

University of Missouri - St. Louis

rathn@umsl.edu

Outline

Introduction

Organic chemistry experiments: Multi-step syntheses, crystallization and structure

determination of organic compounds

Inorganic chemistry experiments: Synthesis, crystallization and structure determination of

transition metal p-toluenesulfonates

Student response

Conclusion

Introduction

Advanced techniques in undergraduate laboratories

Local and regional workforce requires students with expertise in advanced instrumentation and techniques

What can we teach and what can students learn with limited exposure to X-ray diffraction?

The “black box” approach?

The experiments The experiments can be carried out in modules

In the absence of access to X-ray instrumentation, available data sets can be used to provide a good flavor for crystallography using public domain software

The transition metal p-toluenesulfonates are good starting materials, reasonably inexpensive and demonstrate synthetic techniques under inert atmosphere

Criteria for selection of the experiments

The compound synthesis should be doable in a typical senior level laboratory course

The experiments should be easily reproducible

In the absence of a diffractometer, it should be possible to use available data sets for crystallography

A Typical Lab One-hour lecture – “everything you always wanted to know

about crystallography in 50 minutes”

A 4-hour lab period for data collection in batches

A 5-hour lab period with everyone in front of a computer to solve and refine the structure and generate figures and tables

Individual “help time” as needed

August 29, 2012 37

Do you plan on adding X-ray crystallography to your undergraduate teaching curriculum?

a) Yes, within the next year

b) Yes, within the next 1-2 years

c) Yes, within the next 3-5 years

d) Not sure

e) No plans at this time

Audience Poll

Reaction of Vanillin with Acetic Anhydride

O

O

O

O

4-acetoxy-3-methoxybenzaldehyde

O

O

HO

+

OO

O

acetic anhydride

NaOH

H2SO4

Vanillin

Vanyllyl Acetate

Unexpected Reaction ProductIdentfy with X-ray dif f raction

Crystallization

The product can be recrystallized from various solvents including methanol, ethanol and acetone.

The crystals shown are from slow evaporation of an acetone solution.

Crystal Data and Structure Refinement Parameters

Empirical formula C14 H16 O7

Formula weight 296.27

Temperature 100(2) K

Wavelength 0.71073 Å

Crystal system Orthorhombic

Space group Pbca

Unit cell dimensions a = 8.9131(8) Å

b = 7.8741(7) Å

c= 41.266(4) Å

Volume 2896.2(4) Å3

Z 8

Density (calculated) 1.359 Mg/m3

Absorption coefficient 0.110 mm-1

F(000) 1248

Crystal size 0.34 x 0.32 x 0.31 mm3

Theta range for data collection 1.97 to 40.43°.

Index ranges -16≤h≤16, -14≤k≤14, -75≤l≤ 74

Reflections collected 120135

Independent reflections 9191 [R(int) = 0.0329]

Absorption correction Semi-empirical from equivalents

Max. and min. transmission 0.9667 and 0.9637

Refinement method Full-matrix least-squares on F2

Data / restraints / parameters 9191 / 0 / 194

Goodness-of-fit on F2 1.042

Final R indices [I>2sigma(I)] R1 = 0.0437, wR2 = 0.1133

R indices (all data) R1 = 0.0593, wR2 = 0.1218

Largest diff. peak and hole 0.633 and -0.343 e.Å-3

Structure of the tri-acetate

Acid Catalyzed Esterification

O

O

HO

+

OO

O

acetic anhydride

Vanillin

H+/H2SO4

O

O

O

O

O

O

O

Condensation of Citral and Malonic Acid

Crystallization

The compound can be easily crystallized by slow evaporation of methanol, ethanol or acetone solutions.

Crystals shown are from methanol.

Crystal Data and Structure Refinement for the Lactone

Empirical formula C13 H18 O4

Formula weight 238.27

Temperature 100(2) K

Wavelength 0.71073 Å

Crystal system Monoclinic

Space group P21/c

Unit cell dimensions a = 8.6592(5) Å

b = 19.6679(11) Å = 107.049(2)°

c = 7.0735(4) Å

Volume 1151.74(11) Å3

Z 4

Density (calculated) 1.374 Mg/m3

Absorption coefficient 0.101 mm-1

F(000) 512

Crystal size 0.41 x 0.35 x 0.30 mm3

Theta range for data collection 2.07 to 36.59°.

Index ranges -14≤h≤14, -32≤k≤32, -11≤l≤11

Reflections collected 30631

Independent reflections 5644 [R(int) = 0.0242]

Completeness to theta = 36.59° 99.2 %

Absorption correction Semi-empirical from equivalents

Max. and min. transmission 0.9704 and 0.9597

Refinement method Full-matrix least-squares on F2

Data / restraints / parameters 5644 / 0 / 226

Goodness-of-fit on F2 1.045

Final R indices [I>2sigma(I)] R1 = 0.0324, wR2 = 0.0906

R indices (all data) R1 = 0.0394, wR2 = 0.0960

Largest diff. peak and hole 0.482 and -0.197 e.Å-3

Molecular Structure of the Lactone

An “ideal” compound for a crystallography experiment

During the reaction, a fused ring system is produced which is difficult to characterize and predict, at least at the level this experiment is carried out

This compound is hard to characterize by NMR as the 1H-spectrum is featureless

Forms well-formed crystals easily

Student Reports

Students submit a report on synthesis and characterization of the compounds.

They are introduced to mercury and CSD for further analysis of the structure (H-bonding, short contacts, comparison of geometrical parameters with similar structures, etc.).

Vanillin Acetate or Tricyclic Lactone? Both are good experiments for multi-step synthesis

and form good crystals.

The lactone experiment has been used lately for crystallography because the structure is relatively difficult to derive without X-ray.

The lactone experiment needs more time for synthetic steps - well planning needed.

Experiments for Inorganic Laboratory: synthesis and X-ray crystallography The following experiments have been carried out in the

senior inorganic chemistry laboratory course Chem4433 for last three years

Transition metal p-toluenesulfonates Binary salts of Mn(II), Fe(II), Co(II) and Ni(II) can be

synthesized by the reaction of the metal with p-toluenesulfonic acid monohydrate (HOTs.H2O) M + 2HOTs.H2O + 3H2O [M(OH2) 4][OTs] 2.H2O + H2

Monomeric tosylates [M(OTs) 2(DMF) 4] and polymeric tosylates [M(OTs) 2(DMF) 2]n can be crystallized by slowly diffusing (layering) Et2O into a solution of M(OTs) 2 in DMF.

X-ray structure determination Students screen, select and mount their own crystals

Data sets are collected under the supervision of a faculty member

The structure solution and refinement is carried out in a 4-5 hour laboratory period

The instructor and all students have used SHELX-TL software for X-ray structure determination

Students prepare the laboratory reports using the J. Am. Chem. Soc. Communication template. The report includes synthesis and characterization of the compound using X-ray diffraction, as well as other spectroscopic methods.

Structure of mononuclear and polymeric [Tm p-Toluenesulfonates]

Typically one of the two forms are produced: - a mono-nuclear trans-[M(OTs)2(DMF)4] or some of the DMF’s

replaced by water molecules

- A polymeric [M(OTs)2(DMF)2]n

Structure of trans-Fe(OTs)2(DMF)4]

Crystal Data and Structure Refinement for [Fe(OTs2)DMF4] Empirical formula C26 H42 Fe N4 O10 S2

Formula weight 690.61

Temperature 100(2) K

Wavelength 0.71073 Å

Crystal system Monoclinic

Space group P21/n

Unit cell dimensions a = 7.8561(7) Å

b = 24.663(2) Å = 111.155(3)°

c = 8.8289(7) Å

Volume, Z 1595.4(2) Å3 ,2

Density (calculated) 1.438 Mg/m3

Absorption coefficient 0.662 mm-1

F(000) 728

Crystal size 0.33 x 0.15 x 0.11 mm3

Theta range for data collection 2.61 to 34.98°.

Index ranges -12≤h≤12, -36≤k≤39, -14≤l≤8

Reflections collected 43608

Independent reflections 6993 [R(int) = 0.0474]

Absorption correction Semi-empirical from equivalents

Max. and min. transmission 0.9295 and 0.8131

Refinement method Full-matrix least-squares on F2

Data / restraints / parameters 6993 / 0 / 201

Goodness-of-fit on F2 1.021

Final R indices [I>2sigma(I)] R1 = 0.0299, wR2 = 0.0754

R indices (all data) R1 = 0.0407, wR2 = 0.0816

Largest diff. peak and hole 0.513 and -0.337 e.Å-3

Structure of trans-Mn(OTs)2(DMF)2(H2O)2]

Crystal Data and Structure Refinement for [Mn(OTs2)(DMF)2(H2O)2] Empirical formula C20 H32 Mn N2 O10 S2

Formula weight 579.54

Temperature 100(2) K

Wavelength 0.71073 Å

Crystal system Triclinic

Space group P-1

Unit cell dimensions a = 6.3281(5) Å = 90.105(5)°.

b = 12.2134(12) Å = 98.940(4)°.

c = 17.1863(17) Å = 95.606(4)°.

Volume, Z 1305.7(2) Å3 ,2

Density (calculated) 1.474 Mg/m3

Absorption coefficient 0.720 mm-1

F(000) 606

Crystal size 0.51 x 0.13 x 0.08 mm3

Theta range for data collection 1.20 to 33.14°.

Index ranges -9≤h≤9, -18≤k≤18, -23≤l≤26

Reflections collected 41188

Independent reflections 9751 [R(int) = 0.0445]

Absorption correction Semi-empirical from equivalents

Max. and min. transmission 0.9467 and 0.7117

Refinement method Full-matrix least-squares on F2

Data / restraints / parameters 9751 / 6 / 338

Goodness-of-fit on F2 1.034

Final R indices [I>2sigma(I)] R1 = 0.0381, wR2 = 0.0897

R indices (all data) R1 = 0.0639, wR2 = 0.1034

Largest diff. peak and hole 0.601 and -0.394 e.Å-3

Structure of [Mn(OTs)2(DMF) 2]n

Crystal Data and Structure Refinement for [Mn(OTs)2(DMF)2]n Empirical formula C20 H28 Mn N2 O8 S2

Formula weight 543.50

Temperature 100(2) K

Wavelength 0.71073 Å

Crystal system Monoclinic

Space group P21/n

Unit cell dimensions a = 14.8262(8) Å = 90°.

b = 5.2111(3) Å = 107.948(3)°.

c = 16.2554(9) Å = 90°.

Volume, Z 1194.79(12) Å3 ,2

Density (calculated) 1.511 Mg/m3

Absorption coefficient 0.775 mm-1

F(000) 566

Crystal size 0.30 x 0.08 x 0.05 mm3

Theta range for data collection 2.23 to 30.86°.

Index ranges -21≤h≤21, -7≤k≤6, -23≤l≤23

Reflections collected 16816

Independent reflections 3769 [R(int) = 0.0445]

Absorption correction Semi-empirical from equivalents

Max. and min. transmission 0.9645 and 0.8002

Refinement method Full-matrix least-squares on F2

Data / restraints / parameters 3769 / 0 / 154

Goodness-of-fit on F2 1.017

Final R indices [I>2sigma(I)] R1 = 0.0360, wR2 = 0.0862

R indices (all data) R1 = 0.0551, wR2 = 0.0972

Largest diff. peak and hole 0.592 and -0.485 e.Å-3

August 29, 2012 60

Do you plan on adding X-ray crystallography to your undergraduate teaching curriculum?

a) Yes, within the next year

b) Yes, within the next 1-2 years

c) Yes, within the next 3-5 years

d) Not sure

e) No plans at this time

Poll Results

Conclusions The organic laboratory experiments have been

successfully incorporated into our curriculum for the past several years.

Student response has been very enthusiastic (anecdotal and anonymous course evaluations).

These experiments add significant learning to the respective labs.

Acknowledgements Professor Christopher Spilling and students in his

organic laboratory course Chem3643

Professor Stephen Holmes and students in the inorganic laboratory course Chem4433

Undergraduate and high school students who have checked the recrystallization of the samples

NSF-MRI funding for the purchase of X-ray diffractometers

Bruker AXS for a benchtop diffractometer for evaluation

X-ray Diffraction Facility at UMSL

Housed in the custom-designed laboratory space in the Center for Nanoscience.

Current instrumentation includes two single crystal and one powder diffractometers.

August 29, 2012 64

Questions and Answers

Any questions?

Please type any questions you may have for our speakers in the Q&A panel and click Send.

How did we do?

When you exit the webinar, please fill out our evaluation survey to let us know. We appreciate your feedback.

Thank you!

29 August 2012 65

April 30 - Vancouver, BC May 2 - Seattle, WA May 4 - Portland, OR May 7 - Toronto, ON May 9 - Ottawa, ON May 11 - Montreal, QC May 14 - Fremont, CA May 16 - Salt Lake City, UT May 18 - Denver, CO August 6 - Minneapolis, MN August 8 - St. Louis, MO August 10 - Memphis, TN November 5 - Bethesda, MD November 7 - New Brunswick, NJ November 9 - New Haven, CT

One-day seminars celebrating 100 years of analytical X-ray in a city near you 9AM - 12PM: X-ray Fluorescence 1PM - 4PM: X-ray Diffraction Seminars are complimentary Breakfast and lunch are included Proceedings bag available for purchase

Sign up at

www.bruker-axs.com/xteamtour2012

August 29, 2012 29 August 2012

Available at www.bruker-axs.com/webinars_sc-xrd.html

29 August 2012 66 August 29, 2012

X-ray Crystallography On-Demand Webinars

29 August 2012

IMTS (International Manufacturing Technology Show)

Chicago, IL – Sep 10-15

20th Annual Users’ Meeting in Single Crystal X-ray Diffraction Bad Herrenalb, Germany – Sep 23-27

ICCBM 14 (International Conference on the Crystallization

of Biological Macromolecules) Huntsville, AL – Sep 23-28

AOAC (Association of Analytical Communities)

Las Vegas, NV – Sep 30 - Oct 3

SciX / FACSS (Federation of Analytical Chemistry and Spectroscopy Societies) Kansas City, MO – Sep 30 - Oct 4

AAPS (American Association of Pharmaceutical Scientists)

Chicago, IL – Oct 14-18

AVS (Science and Technology of Materials, Interfaces, and Processing) Tampa, FL – Oct 28 - Nov 2

MRS Fall (Materials Research Society)

Boston, MA – Nov 26-30

29 August 2012 67

Upcoming Events Come Visit Us!

August 29, 2012 29 August 2012

29 August 2012 68

Like what you learned in this webinar?

Subscribe to Bruker’s FIRST Newsletter to get webinar announcements, fascinating articles, and analytical X-ray news delivered right to your inbox!

Subscribe at: www.bruker-axs.com/bruker_axs_newsletter.html

August 29, 2012 29 August 2012

Thank you for attending!

www.bruker.com

29 August 2012 69