Answer Lab questions

Synthesis and Characterisation
Techniques in Chemistry
CHEM0024/6 LABORATORY
MANUAL
2022/2023
Student Name:
Laboratory Coordinators:
Dr Derek Macmillan
(d.macmillan@ucl.ac.uk)
Dr Helen Allan
(h.allan@ucl.ac.uk)
TABLE OF CONTENTS
SAFETY
1
CONDUCT IN THE LABORATORIES ……………………………………………………………………………………………. 1
ACCIDENTS ……………………………………………………………………………………………………………………. 1
FIRE ALARMS………………………………………………………………………………………………………………….. 1
SAFETY ASSESSMENTS (RISK & COSHH ASSESSMENTS) ………………………………………………………………….. 2
SAFETY TRAINING AND RISK ASSESSMENT IN THE UNDERGRADUATE LABORATORIES ………………………………………….. 2
RISK ASSESSMENT ………………………………………………………………………………………………………………………. 2
COSHH ASSESSMENT………………………………………………………………………………………………………………….. 2
WASTE DISPOSAL PROCEDURES …………………………………………………………………………………………….. 5
1) SOLID CHEMICAL WASTE ……………………………………………………………………………………………………………. 5
2) LIQUID CHEMICAL WASTE ………………………………………………………………………………………………………….. 5
3) DAMAGED/BROKEN GLASSWARE …………………………………………………………………………………………………. 5
4) SHARPS ……………………………………………………………………………………………………………………………….. 5
5) GENERAL LABORATORY WASTE …………………………………………………………………………………………………… 5
GENERAL LABORATORY INSTRUCTIONS
6
GOOD LABORATORY PRACTICE………………………………………………………………………………………………. 6
BENCHES AND APPARATUS ………………………………………………………………………………………………….. 6
ISSUE OF APPARATUS AND CHEMICALS …………………………………………………………………………………….. 6
PHYSICAL APPARATUS AND SPECTROMETERS………………………………………………………………………………. 6
CHEM0024/6 OVERVIEW
7
OBJECTIVES ……………………………………………………………………………………………………………………. 7
COURSE REQUIREMENTS …………………………………………………………………………………………………….. 7
ASSESSMENT ………………………………………………………………………………………………………………….. 8
FEEDBACK ……………………………………………………………………………………………………………………… 8
KEEPING A GOOD LABORATORY NOTEBOOK (TASK 4) ……………………………………………………………………. 10
POST-LAB SUBMISSIONS …………………………………………………………………………………………………… 14
FINALLY………………………………………………………………………………………………………………………. 16
PRACTICAL A: REACTION OF FERROCENE COMPOUNDS
17
LEARNING OBJECTIVE ………………………………………………………………………………………………………. 17
RELATES TO COURSE MATERIAL …………………………………………………………………………………………… 17
INTRODUCTION ……………………………………………………………………………………………………………… 17
OBJECTIVES ………………………………………………………………………………………………………………….. 20
PRE-LAB (REVISIT P10-P13 FOR DETAILS) ………………………………………………………………………………… 20
PRACTICAL …………………………………………………………………………………………………………………… 21
IMPORTANT NOTES …………………………………………………………………………………………………………………… 21
PART 1. OXIDATION OF FERROCENE TO FERROCENIUM HEXAFLUOROPHOSPHATE ………………………………………….. 21
PART 2. C-H ACTIVATION AND HECK REACTION …………………………………………………………………………………. 21
RESULTS ……………………………………………………………………………………………………………………… 24
REFERENCES …………………………………………………………………………………………………………………. 24
PRACTICAL A MARK SCHEME (TASKS 5 AND 6) ………………………………………………………………………….. 25
PRACTICAL B: REACTIONS OF CARBOHYDRATES AND COMPLEX NMR
26
LEARNING OBJECTIVE ………………………………………………………………………………………………………. 26
RELATES TO COURSE MATERIAL …………………………………………………………………………………………… 26
INTRODUCTION ……………………………………………………………………………………………………………… 26
CARBOHYDRATES ……………………………………………………………………………………………………………………… 26
RADICAL BROMINATION SOLVENTS …………………………………………………………………………………………………. 27
COMPLEX NMR COSY/DEPT ……………………………………………………………………………………………………… 28
OBJECTIVES ………………………………………………………………………………………………………………….. 30
PRE-LAB (REVISIT P10-P13 FOR DETAILS) ………………………………………………………………………………… 30
REFERENCE SPECTRA ……………………………………………………………………………………………………….. 31
PRACTICAL …………………………………………………………………………………………………………………… 36
IMPORTANT NOTES …………………………………………………………………………………………………………………… 36
1. BENZYLIDENE PROTECTION OF METHYL-Α-D-GLUCOPYRANOSIDE ………………………………………………………….. 36
2. RADICAL BROMINATION ………………………………………………………………………………………………………….. 36
RESULTS ……………………………………………………………………………………………………………………… 38
REFERENCES …………………………………………………………………………………………………………………. 38
PRACTICAL B MARK SCHEME (TASKS 5 AND 6) ………………………………………………………………………….. 39
PRACTICAL C: SYNTHESIS OF CRYSTALLINE FUNCTIONAL MATERIALS
40
LEARNING OBJECTIVE ………………………………………………………………………………………………………. 40
RELATES TO COURSE MATERIAL …………………………………………………………………………………………… 40
INTRODUCTION ……………………………………………………………………………………………………………… 40
HYDROXYAPATITE …………………………………………………………………………………………………………………….. 41
LITHIUM IRON PHOSPHATE ………………………………………………………………………………………………… 42
OBJECTIVES ………………………………………………………………………………………………………………….. 43
PRE-LAB TASKS………………………………………………………………………………………………………………………… 43
TASK 1. SAFETY ASSESSMENT ……………………………………………………………………………………………………….. 43
TASK 2. TABLE OF REAGENTS ……………………………………………………………………………………………………….. 43
TASK 3. PRE-LAB QUESTIONS……………………………………………………………………………………………………….. 43
TASK 4. LAB NOTEBOOK……………………………………………………………………………………………………………… 44
PRACTICAL …………………………………………………………………………………………………………………… 44
SCHEDULE………………………………………………………………………………………………………………………………. 44
IMPORTANT NOTES …………………………………………………………………………………………………………. 45
DAY 1: ………………………………………………………………………………………………………………………………….. 45
HA SYNTHESIS …………………………………………………………………………………………………………………………. 45
LFP SYNTHESIS ………………………………………………………………………………………………………………………… 47
DAY 2 …………………………………………………………………………………………………………………………………… 48
POST-LAB TASKS ……………………………………………………………………………………………………………. 48
TASK 5. JUSTIFICATION OF YIELD AND PURITY ……………………………………………………………………………. 48
TASK 6. EXPERIMENTAL EVALUATION …………………………………………………………………………………….. 49
FINAL REPORT QUESTIONS…………………………………………………………………………………………………. 49
REFERENCES …………………………………………………………………………………………………………………. 52
PRACTICAL C MARK SCHEME (TASKS 5 AND 6) ………………………………………………………………………….. 53
PRACTICAL D: PEPTIDE SYNTHESIS (ORGANIC NATSCI AND MEDCHEM ONLY)
54
LEARNING OBJECTIVE ………………………………………………………………………………………………………. 54
RELATES TO COURSE MATERIAL …………………………………………………………………………………………… 54
INTRODUCTION ……………………………………………………………………………………………………………… 54
OBJECTIVES ………………………………………………………………………………………………………………….. 57
PRE-LAB (REVISIT P10-P13 FOR DETAILS) ………………………………………………………………………………… 57
REFERENCE SPECTRA ……………………………………………………………………………………………………….. 58
PRACTICAL …………………………………………………………………………………………………………………… 62
IMPORTANT NOTES …………………………………………………………………………………………………………………… 62
1. PREPARING THE RESIN AND REMOVING FMOC ………………………………………………………………………………… 62
2. GLYCINE COUPLING PROCEDURE ………………………………………………………………………………………………… 63
3. FMOC DEPROTECTION …………………………………………………………………………………………………………….. 63
4. ALANINE COUPLING/FMOC DEPROTECTION……………………………………………………………………………………. 63
5. GLYCINE COUPLING/FMOC DEPROTECTION ……………………………………………………………………………………. 63
6. ALKYL CHAIN COUPLING…………………………………………………………………………………………………………… 63
7. PEPTIDE CLEAVAGE ………………………………………………………………………………………………………………… 63
TLC ANALYSIS …………………………………………………………………………………………………………………………. 64
SELF-ASSEMBLY AND FTIR ANALYSIS ………………………………………………………………………………………………. 64
RESULTS ……………………………………………………………………………………………………………………… 64
REFERENCES …………………………………………………………………………………………………………………. 65
PRACTICAL D MARK SCHEME (TASKS 5 AND 6) ………………………………………………………………………….. 66
APPENDICES
67
APPENDIX 1 …………………………………………………………………………………………………………………. 67
HAZARD STATEMENTS ……………………………………………………………………………………………………………….. 67
PRECAUTION STATEMENTS ………………………………………………………………………………………………………….. 68
APPENDIX 2 …………………………………………………………………………………………………………………. 72
MAKING UP A SAMPLE FOR NMR ………………………………………………………………………………………………….. 72
APPENDIX 3 …………………………………………………………………………………………………………………. 73
THE EVANS METHOD FOR DETERMINING MAGNETIC SUSCEPTIBILITIES IN SOLUTION ……………………………………… 73
APPENDIX 4 …………………………………………………………………………………………………………………. 75
APPENDIX 5 …………………………………………………………………………………………………………………. 76
FAQS……………………………………………………………………………………………………………………………………. 76
SAFETY
Conduct in the Laboratories
The following pages present some guiding principles for safety. All persons engaged in work within the
department have agreed to abide by both UCL statutory requirements and departmental safety rules.
Laboratories are potentially dangerous workplaces and accidents in the lab can have serious and tragic
consequences. However, if you are aware of potential hazards, and work with due care and attention
to safety, the risk of accidents is small.
•
•
Always work carefully and tidily, use your common sense, and abide by the safety regulations as
presented in the departmental safety handbook.
Your own safety, and that of your colleagues in the lab, is largely determined by your work
practices.
The laboratory is a place for carrying out practical work. Please confine your other activities to more
suitable places.
•
•
•
•
•
•
•
•
You must sign in to laboratory sessions (electronically, tap-in using your UCL ID on the
Register@ucl system). You must arrive at the laboratory between 09:00 and 09:30 am.
Eye protection & lab coats MUST be worn at all times.
Eating and drinking are banned in laboratories.
Sensible shoes and clothing must be worn (skin on legs and feet must be covered), material should
preferably be made of natural fibre (cotton, wool etc.).
Bags and coats should be stored in lockers, not brought to the laboratory, unless instructed to do
so.
It is acceptable to use your mobile phone for calculations, accessing UCL intranet or camera
functions. However, personal calls etc. should be made outside the laboratory.
Some of the materials that you will use will require the use of appropriate gloves. The information
for this will come from the COSHH and risk assessments for the experiment. Disposable gloves
must be changed on a regular basis or if they become soiled.
You should wash your hands and remove safety glasses and lab coats at the end of the session
before leaving the laboratory.
Accidents
It is vital to know what to do in case of an accident. All accidents or undesired incidents no matter how
slight must be reported to a demonstrator/member of staff who will assist with first aid and give advice
on reporting the incident. Reporting of accidents is a UCL requirement and the information obtained
will be used to help to prevent future/more serious incidents occurring.
Fire Alarms
If you are in a practical class when the alarms sound, spend a few seconds turning off heating
equipment or making your experiment safe, then leave the building by the nearest marked exit. Do
not attempt to enter the building until told it is safe to do so.
CHEM0024 Lab Manual |1
Safety Assessments (Risk & COSHH assessments)
As undergraduates you will be required to perform practicals to give you experience of handling
reagents, practical techniques, manipulating glassware, using electrical equipment and the fumehood
facilities available in the laboratories.
Risk and COSHH assessments are required to be completed before any activity is undertaken. The staff
supervising laboratory classes will have undertaken risk & COSHH assessments for the practicals you
will perform; these are available on Moodle or a hard copy is available in the laboratory. You are
required to carry out and record your own safety assessments and enter them in your notebooks for
inspection before beginning any practical work.
Safety training and risk assessment in the undergraduate laboratories
The department has a structured approach to safety training and risk assessment. The goal is to be
able to understand and write your own safety assessments for practical chemistry, this will be an
essential skill to have for advanced practicals in the 3rd year and research projects in the 4th year.
You must look at the UCL Department of Chemistry Safety page on Moodle and complete the risk
assessment lesson. You will also need to complete the risk assessment and pass the pre-lab safety
quiz before you start each practical (on the Synthesis 1 Moodle page).
Risk Assessment
Risk assessment is the process by which you think through a procedure to minimize the chances of
things going wrong. It involves planning the activity, deciding what could go wrong, what harm could
be caused and to whom, how likely this is, and determining the precautions necessary to prevent the
harm. A risk assessment is a combination of COSHH and identifiable hazards of the activity taking place.
You must read through the practical and think about each operation you will carry out and consider
what could go wrong, how and why. You need to think about what can be done to prevent this from
happening (precautions) and what to do should this occur (treatment).
There are 5 steps to risk assessment:
•
•
•
•
•
Identify hazards
Assess their likelihood
Control the risk
Record
Revise (useful to know but not required by undergraduates)
A risk assessment looks at the whole process that is going on, COSHH will give the chemical hazards,
but there are still the hazards of the actual process, which you must consider.
COSHH Assessment
COSHH stands for the Control Of Substances Hazardous to Health. A COSHH assessment involves
finding out what the health hazards of chemicals are, and deciding how to prevent harm to health.
Read the practical procedure and look at the chemicals involved. Using the Sigma Aldrich website you
can look up and find the hazards/dangers of each chemical and decide how best to minimise these
hazards (the safety data sheet (SDS) always provides precautions for use). A list of H (Hazard) and P
(Precaution) phrases can be found in appendix 1 at the end of the lab manual.
2|CHEM0024 Lab Manual
Click
on
the
Safety
&
Documentation tab or scroll down
Also take time to read through
the Safety Data Sheet (SDS)
Hazard and precaution statements
should be copied into your lab book
(not just the numbers) – see
appendix for list of phrases
Example risk assessment: Borohydride reduction of a ketone
Task
Name of Practical – Borohydride reduction of a ketone
Location –Turner Lab
Description of practical – Reduction of a ketone using sodium borohydride in methanol/water at 0
°C. TLC (silica plate) run using ethyl acetate/pet. ether. Glass TLC spotters made using a Bunsen
burner. Reaction worked up by saturating with sodium chloride and extracting with diethyl ether
using a separating funnel. Dried using magnesium sulfate. Rotary evaporator used. Product purified
by distillation. IR spectrum recorded.
Persons at Risk – Students, staff, visitors and contractors
Hazards; A list of all equipment (physical), substances or actions (procedures) which are
hazardous.
Physical Hazards
• Handling of chemicals (spills, incorrect disposal) – can cause slips/trips/chemical burns or fires.
• Electrical equipment (hotplates, balances) – potential for electric shock/ burns or fire
• Inappropriate or rough use of glassware may result in cuts and injury
• Making TLC spotters – risk of burns from hot glass/ Bunsen burner, cuts and puncture wounds
from glass spotters.
• Use of separating funnel – potential for pressure build up
Chemical Hazards
• Ketone – Flammable
• Sodium borohydride – In contact with water releases flammable gas which may ignite
spontaneously, toxic if swallowed, causes severe skin burns and eye damage, may damage
fertility or unborn child.
CHEM0024 Lab Manual |3
•
•
•
•
•
Methanol – Highly flammable, toxic if swallowed, in contact with skin or inhaled.
Ethyl acetate – Highly flammable, causes serious eye irritation, may cause drowsiness.
Petroleum ether – Highly flammable, harmful if swallowed, causes skin irritation, may cause
drowsiness, toxic to aquatics.
Diethyl ether – Extremely flammable, harmful if swallowed, may cause drowsiness.
Magnesium sulfate – No known hazard.
Controls Measures; Put in place to prevent the risk occurring
Physical Controls
• Check all equipment before using, do not touch electrical equipment with wet hands.
• Handle glassware with care, clean up breakages immediately with a dustpan and brush and
dispose of in the yellow glass bin by stores. Inform a staff member.
• Ensure that there are no flammable chemicals in the vicinity of the Bunsen burners.
• Dispose of TLC spotters immediately after use in the glass bin do not leave on bench tops.
• When using a separating funnel vent regularly to avoid the build-up of gas.
Hazardous Chemical Controls
• Keep flammable chemicals away from heat sources and open flames. Never heat on a hot plate.
• Avoid breathing fumes, if inhaled remove to fresh air.
• Dispose of ethyl acetate, pet. ether and diethyl ether into the non-chlorinated waste container
opposite stores. Magnesium sulfate should be disposed of into the solid waste container.
General Safety Controls;
•
•
•
•
Spills should be cleaned up as soon as possible and/or reported.
PPE (Personal Protective Equipment) to be worn at all times – including laboratory coat and
safety goggles.
Unless stated by the Safety Data Sheet, chemicals in contact with skin and eyes should be
thoroughly washed with water. (Eye wash stations for eyes).
Training is provided on the use of all instruments, chemicals and appropriate storage / disposal.
Ask a demonstrator if you are unsure.
Residual Risk; This is the risk remaining after the controls above are applied
Low (after implementation of control measures.)
For each practical the risk assessment, minus the chemical hazards and hazardous chemical controls,
are given to you on Moodle. You are expected to look up the COSHH assessments for the chemicals
that you will be using and fill the chemical hazard and chemical controls sections in. You should then
print your risk assessment and stick it in your lab book. You must also complete the pre-lab safety quiz
on Moodle before the start of each practical.
Before you are allowed to commence any lab work you must have completed the Moodle pre-lab
quiz. Your COSHH and risk assessment must also be completed and will be assessed in the laboratory
by lab demonstrators on the 1st day of each practical.
4|CHEM0024 Lab Manual
Waste Disposal Procedures
The disposal of waste from the laboratory is split into 5 streams. Each of these streams has its own
disposal procedures, which are a direct consequence of the hazards associated with the materials, you
will be advised as to which stream to follow.
At no time should noxious substances be disposed of down any sink
1) Solid chemical waste
Different labelled containers are provided for solid waste,
which may include the following:
•
•
•
Chromatography Waste (Silica or alumina)
Solid reagents
Contaminated paper e.g. filter paper
2) Liquid chemical waste
Different labelled waste bins are provided for liquid
waste, which may include the following:
•
•
•
•
Chlorinated Solvents
Non Chlorinated Solvents
Heavy metal waste solutions
Mercury metal
3) Damaged/broken Glassware
All glassware must be disposed of in the yellow plastic bins provided; under
no circumstances should broken glass enter the general laboratory waste
stream. Glass pipettes should go in this bin.
4) Sharps
Sharps are a general term for syringe needles and cannulas. Needles and Cannulas
for disposal must be placed in the yellow ‘sharpsafe’ bins. Under no circumstances
should these items enter the general waste stream.
5) General Laboratory Waste
All other uncontaminated laboratory waste (gloves, blue roll etc.) should be
placed in the general yellow bag laboratory waste bins.
CHEM0024 Lab Manual |5
GENERAL LABORATORY INSTRUCTIONS
Good Laboratory Practice
It is imperative that all members of the class contribute to the smooth running of the laboratory and
assist the technical staff in maintaining a clean, safe and tidy working environment. To that end all
students must make sure that their work area is clean and tidy before they leave the laboratory.
Chemicals must not be left beside the balances or on the benches but must be returned to the front
bench or side racks. It is essential that when working in the spectroscopy laboratory you tidy the bench
tops when your measurements are complete. Surfaces must not be left contaminated with discarded
pipettes or other debris. Spillages and broken glass must be dealt with immediately; if that is not done
then you endanger the health of all members of the laboratory.
Benches and Apparatus
You have been allocated a box kit which you will work from for the whole of this lab course. The kit
contains a set of basic apparatus required for the experiments you will undertake. Box kits stored are
on shelves located at the back of the lab and should be returned to these shelves at the end of each
lab session. You will be allocated suitable bench space to work on for a given session, keep in mind
where fixed equipment for the practical you are undertaking is located. All equipment you need should
be allocated on your bench, or nearby (see detailed lab plan on Moodle). Clamps and bosses are found
in the central access cupboards 34 and 35. Water bottles will be on the bench tops. Do not leave
apparatus on the benches overnight unless you have special permission to do so. Bench tops must be
cleaned at the end of each day.
Issue of Apparatus and Chemicals
A set of common reagents are provided. The laboratory staff will ensure that these bottles are kept
filled. A comprehensive collection of other reagents is kept in the designated area for that experiment
– check the list shown in the lab. These are for general use, so be careful to avoid contaminating them.
Under no circumstances may they be taken to your bench. Do not take more reagent than you need
and do not return unused portions to the bottle. Other reagents and equipment will be issued from
the stores as required. If you cannot find a particular piece of apparatus check the list shown in the lab
and see whether it is issued from stores. Volatile and toxic liquids will be dispensed using Dispensette
dispensers.
Physical Apparatus and Spectrometers
A number of precise physical instruments will be used during this course; these are sensitive and
delicate and must be used with great care. In particular, chemicals must not be allowed to come into
contact with them. A range of NMR and infrared spectrometers are available for use in this laboratory
course.
If you are uncertain about the use of any piece of apparatus consult with one of the demonstrators or
the laboratory staff.
6|CHEM0024 Lab Manual
CHEM0024/6 OVERVIEW
Objectives
The course will build on 1st and 2nd year practical laboratory courses by developing training in more
advanced practical skills, experimental techniques and spectral analysis. The course will provide a firm
foundation for pursuing a 4th year research project (CHEM0080) with a high level of independence.
•
•
•
•
•
•
•
•
To be able to conduct a risk assessment in addition to the COSHH assessment and apply suitable
safety measures for the proposed practical work.
The ability to organise time to plan experiments and complete them in an allotted time safely and
successfully.
Identify/Select appropriate equipment for a chemical reaction.
Maintain a laboratory notebook to a professional standard.
Apply core chemical reactions to the preparation of compounds.
Design and conduct experiments to interrogate a research hypothesis or problem.
Identify problems/troubleshoot reactions during/afterwards.
Critically interrogate results and spectral data to assess the outcome of the practical.
Course Requirements
The course consists of three practicals, each practical will take 2 days – you will be assigned a practical
order. In these practicals you will:
•
•
•
•
Oxidise ferrocene and perform a palladium-catalysed Heck reaction
Apply protecting group strategy and radical chemistry to synthesise a substituted carbohydrate
(Not Inorganic NatSci)
Functional material synthesis (not Organic NatSci or MedChem, Inorganic NatSci do two of these
syntheses)
Synthesise a short peptide using solid phase synthesis (Organic NatSci and MedChem only)
In each practical different isolation, purification and analytical techniques are used (e.g. workup/partition exploiting the differences in solubility of different reagents, recrystallization, column
chromatography & distillation, and analysis by IR, NMR & TLC). You will use these techniques to isolate
compounds and gain important data on them.
CHEM0024 Lab Manual |7
Assessment
The course is assessed via a combination of completed lab-based tasks, your lab book, analytical data
you collect, and one final “formal” report according to the explicit criteria described below. It is in your
interest to ensure your results are consistent with any predictions you may make, or if they are not
consistent they are repeatable and can be rationalised.
The following Tasks are completed either before or during the lab.
1.
2.
3.
4.
5.
6.
A safety assessment.
Calculations of quantities of reagents, and accurate reaction equations.
Pre-lab questions relating to understanding of the reaction.
Keeping a good lab book (how repeatable is the work you have performed?)
Laboratory skills judged by yield and purity
Briefly explain how you know the experiment has worked?
The Pre-lab (Tasks 1-3) must be completed before attending the lab. Tasks 1 and 2 may both be
marked in the lab by laboratory demonstrators on the first day of each practical.
Tasks 3-6 will be marked by submission to Moodle. After each practical, scans of the relevant pages
in your lab book, including Tasks 1 and 2, must be submitted via Moodle by the appropriate deadline.
Each practical you submit will be worth 25% of the marks available for Synthesis 1. At the end of
synthesis 1 you will be allocated one practical ”write-up” for which you should complete a short, but
more detailed, formal piece of writing, in the style typical of a research paper or “supporting
information”. Further details will be provided in due course.
The overall aim is to assess the skills you have developed in keeping your lab book and in planning and
monitoring reactions, as well as how you judge if a reaction has been successful from the interpretation
of the physical data.
In recognition of the fact that experimental work is an essential part of chemical education, (in the
absence of an authorised Extenuating Circumstance), you must attend 100% of your lab sessions and
submit all prelab work and final reports.
Feedback
Each experiment will be marked either in the laboratory (Tasks 1-2) or via Moodle (Tasks 3-6) and
feedback will be provided. Students are encouraged to discuss and obtain feedback with the
demonstrators in the lab.
Any marking queries should have VERY STRONG justification and may result in marks going down.
Practicals A, B and D: These can only be discussed during Dr Macmillan’s office hours – emailed marking
queries will not be answered.
Practicals C and C2: Please contact Dr Yang Xu.
8|CHEM0024 Lab Manual
A guide to assessment and feedback. A general marking scheme for Tasks 1-4 for all practical experiments is described below:
Task
1.
Assessment
Safety
Not Attempted/0
Emerging/1
Developing/2
Satisfactory/3
Exceptional/4
Not
present
or
completely irrelevant
Some hazards are
identified, but the
majority are missing.
All critical hazards are
identified. -AND EITHERAll hazards are present for
most reagents. -OR- Most
hazards are present for all
reagents. In either case 13 may be missing
All hazards are clearly
identified and included
for all reagents.
As well as hazards generally true of
the reagent, the assessment takes
into account safety measures taken,
likelihood of risk and scale to
produce a thorough consideration
of risk
A critical hazard (e.g.
pyrophoric) is missing.
Task 2.
Reagents
Table
of
Not
present
or
completely irrelevant
Some reagents are
missing. -OR- There are
multiple minor errors. OR- There is a major
error.
All reagents are present
but there is a minor error.
The amounts (moles and
mass) of all reagents are
accurately calculated.
As well as moles and mass, liquid
reagents have been identified and
volumes have been calculated.
Reaction equations are complete
and correct.
Task 3. Prelab Questions
(Understanding
of
reaction)
Not
present
or
completely irrelevant
Answers
are
incomplete. (Either the
answers are incomplete
or not all questions are
answered).
Provided
insufficient
support/evidence/
Addressed all of
chemistry/method/
Addressed all the
chemistry/method/
explanation
for
comments
about
chemistry/method/
interpretation
with
evidence and support, but
with occasional lack of
clarity or with minor
error. -OR- Good answer
to
a
slightly
misinterpreted question.
interpretation in sufficient detail,
with evidence/support, and good
clarity.
The
procedure
is
sufficiently detailed to
repeat the experiment,
some observations are
included.
The notebook is clearly presented,
including thorough observations
and sufficient procedural detail for
another chemist to repeat the
experiment.
Interpretation or missing
some key ideas. -ORGood answer to a totally
misinterpreted question.
Task 4. Lab Notebook
Not present, illegible or
completely irrelevant
Key information is
missing
from
the
experimental
procedure.
The
procedure
is
sufficiently detailed to
repeat the experiment,
but no observations are
included.
the
points
of
CHEM0024 Lab Manual |9
Keeping a good laboratory notebook (Task 4)
It is an essential part of your training that you learn to keep a good laboratory notebook. However,
your laboratory notebook is a working document. This means it does not have to be perfectly
presented, but needs to be legible. It is somewhere for you to make calculations, record weighing and
measurement values and make notes useful to you. The acid test is that someone else could repeat
the practical and get the exact same results from your information. As such you should not record
notes anywhere else (in the lab script or on other pieces of paper). You will not be assessed on how
‘tidy’ your notebook is as long as it is legible.
Your notebook must contain an accurate primary record of every practical you carry out. Thus, you
must record everything, i.e. your procedures, calculation of masses, observations (such as temperature
change, colour change, precipitation, gas evolution, etc.), weights, and so on directly into your lab
book. Omissions of key facts or observations not noted by the end of the lab may be penalised. Write
directly into the notebook and not any other piece of paper.
Make sure your name is on the front and/or the inside cover of your lab book and number the pages.
Leave one or two pages free at the beginning for an index.
You should aim to complete all lab work before 5:30pm on the 2nd day, such that demonstrators can
check that you have recorded all weights, measurements, observations, and calculations, and can
sign it off as being complete.
Present your practical work using the following format:
(1) Task 1: Print your completed Hazard and Risk assessment (COSHH assessment) for the reaction
and stick this in your lab book. This is a vital (and legally required) safety measure, RE-READ it
CAREFULLY and make sure that you understand why the precautions are being used and comply
with them.
(2) Task 2: Write down the date and the title of the practical. Next you must have the stoichiometric
equation for each of the reactions you are going to carry out. Indicate the reagents, solvents and
conditions that you will use above the arrow leading to the product. Give the product a unique
code number constructed from your initials and the page number on which the practical is being
written up. Thus, if A W Williamson is recording work on page 39 of their lab book, their desired
product should be given the code number AWW-39. This code number should then appear on the
sample label and on all the spectra obtained from the sample (as well as the formula and structure
of the sample). If more than one product is isolated they can be labelled AWW-39a, AWW-39b
etc.
Beneath the reaction equation, tabulate the relative molecular masses of the reagents, the
masses used in the practical and the corresponding molar quantities. Identify the limiting reagent
and show the theoretical yield (including working out how you got to this value – it is useful to do
this on the graph paper side).
10 | C H E M 0 0 2 4 L a b M a n u a l
(3) Task 3: Complete the additional required Pre-Lab exercises.
(4) Task 4: A concise account of the essential details of your procedure should be present. Each new
day’s work should be dated in the margin. There should be sufficient information that a
competent chemist could reproduce your results exactly. Thus quantities of reagents,
temperatures, periods of time for procedures, methods of product isolation and purification etc.
are all important information, and must always be recorded. However, it is not necessary to
describe in detail or draw diagrams of the equipment since only standard set-ups are used. It is
often useful to put in your rough notes the time when you do something particularly when it is
the start of an operation, such as heating at reflux or a dropwise addition that needs to be timed.
TLC plates should be copied into your lab book in an appropriate place and Rf values calculated.
(5) Task 5: The yield (i.e. weight of product isolated) should be recorded and the actual percentage
yield (yield divided by theoretical yield multiplied by 100) calculated to the nearest percentage
and noted in brackets. Any practical measurements you have made should be included and all
spectra labelled with your sample number, your assigned chemical formula and the structure of
the compound, and stuck into your lab book. It is useful to record yields and physical data for both
crude and purified materials.
(6) Task 6: Include a brief description of how you know your reaction has worked. Guidance for
completion of this task is available in Appendix 4.
See below for example lab book pages.
C H E M 0 0 2 4 L a b M a n u a l | 11
Example of a lab book entry (Tasks 1-3)
Name of Practical – Benzyl protection of 6-hydroxybenzothiazole
Location –Turner Lab
Description of practical – Reaction of 6-hydroxybenzothiazole with benzyl bromide using potassium
carbonate in acetone. TLC (silica plate) run using ethyl acetate/pet. ether. Glass TLC spotters made
using a Bunsen burner. Reaction worked up by filtration and concentrated using a rotary evaporator.
Product purified by column chromatography using ethyl acetate/pet. ether. IR and H NMR spectra
recorded.
Persons at Risk – Students, staff, visitors and contractors
Hazards; A list of all equipment (physical), substances or actions (procedures) which are
hazardous.
Physical Hazards
• Handling of chemicals (spills, incorrect disposal) – can cause slips/trips/chemical burns or fires.
• Electrical equipment (hotplates, balances) – potential for electric shock/ burns or fire.
• Inappropriate or rough use of glassware may result in cuts and injury.
• Use of rotary evaporator – Moving parts could trap clothes or hair, damaged glassware could
break under vacuum causing injury.
• Use of column chromatography – potential for pressure build up, use of too strong a flow of
compressed air may cause adaptor to blow out and break.
Chemical Hazards
• 6-hydroxybenzothiazole – Harmful if swallowed, causes severe eye irritation.
• Benzyl bromide – Lachrymator, causes serious skin and eye irritation.
• Potassium carbonate – Harmful if swallowed, causes severe eye and skin irritation.
• Acetone – Highly flammable, causes serious eye irritation, may cause drowsiness.
• Ethyl acetate – Highly flammable, causes serious eye irritation, may cause drowsiness.
• Petroleum ether – Highly flammable, harmful if swallowed, causes skin irritation, may cause
drowsiness, toxic to aquatics.
• Silica – Causes serious lung irritation.
Controls Measures; Put in place to prevent the risk occurring
Physical Controls
• Visually inspect hotplates and glassware before using (check for exposed wires or melted cables
or star cracks in glassware). Do not touch electrical equipment with wet hands.
• Handle glassware with care, clean up breakages immediately with a dustpan and brush and
dispose of in the yellow glass bin by stores. Inform a staff member.
• When using a rotary evaporator check glassware for cracks before applying vacuum. Tie back
hair. Do not attempt to stop the rotation using anything other than the controls.
• When performing column chromatography take care when transporting kits. Ensure
compressed air is used with reason and slow increase of pressure used.
Hazardous Chemical Controls
• Keep flammable chemicals away from heat sources and open flames. Never heat on a hot plate.
• Avoid breathing fumes, if inhaled remove to fresh air.
• Dry silica should only be used in the fumehood.
• Dispose of acetone, ethyl acetate and petroleum ether into the non-chlorinated waste
container opposite stores. Silica should be disposed of into the solid waste container.
12 | C H E M 0 0 2 4 L a b M a n u a l
General Safety Controls;
•
•
•
•
•
Spills should be cleaned up as soon as possible and/ or reported.
PPE (Personal Protective Equipment) to be worn at all times – including laboratory coat and
safety goggles.
Unless stated by the Safety Data Sheet, chemicals in contact with skin and eyes should be
thoroughly washed with water. (Eye wash stations for eyes).
Wash hands before taking any breaks and at the end of work.
Training is provided on the use of all instruments, chemicals and appropriate storage / disposal.
Ask a demonstrator if you are unsure.
Residual Risk; This is the risk remaining after the controls above are applied
Low (after implementation of control measures.)
COMPONENT
C H E M 0 0 2 4 L a b M a n u a l | 13
Post-Lab Submissions
You must submit scans of the appropriate pages of your lab book via the submission link in Moodle by
the appropriate deadline.
Scanning is easily achieved using the Print@UCL printers. After accessing the printer menu select the
‘Email’ icon.
From the options select the ‘Layout Adjustment’ tab.
Within these options select ‘Original Size’.
From these options select ‘Preset Scan Area’.
14 | C H E M 0 0 2 4 L a b M a n u a l
Select the first entry for ‘A4’ (A4 portrait orientation) and press ‘OK’.
Then select the Job Assembly tab and press ‘Build Job’.
Select ‘On’ and press ‘OK’.
C H E M 0 0 2 4 L a b M a n u a l | 15
Then scan your lab book a page at a time by pressing the large green physical button on the printer by
positioning your lab book as shown below.
Once finished select ‘Submit Job’ to send a .pdf of your scans to your email address. Please change the
name of the file you receive to format SURNAME_PRACTICAL NUMBER.pdf before submitting via
Moodle. Please check that your scan is complete before uploading your submission.
On rare occasions there can be genuine submission problems associated with Moodle. Late reports
will be treated as such, and appropriate penalties applied, unless you can produce an ISD ticket receipt
detailing the nature of the problem and how (and when) it was resolved. Any technical issues that are
not accompanied with an ISD enquiry receipt will be dismissed.
Finally
Samples must be handed in to the stores at the end of your final laboratory class.
16 | C H E M 0 0 2 4 L a b M a n u a l
PRACTICAL A: REACTION OF FERROCENE
COMPOUNDS
Learning Objective
To study the oxidation of the iron centre in ferrocene and determine the magnetic moment of its
oxidation product. To synthesise a planar chiral derivative of ferrocene using a dehydrogenative Heck
reaction. To understand the different reaction conditions required for these transformations and also
why certain methods/techniques are used to isolate and purify products. To be able to predict key
differences in the NMR spectra of starting materials and the products and assign the spectra of the
products.
Relates To Course Material
•
•
•
Bonding in ferrocene and magnetic moments of transition metals CHEM0014 (FC lecture notes)
Electrophilic substitution CHEM0016 (BS lecture notes)
Heck reaction CHEM0034 (JCA lecture notes)
Introduction
Organometallic chemistry is at the interface of organic chemistry and the chemistry of the
metals, since an organometallic compound is defined as one containing a metal-to-carbon
bond. Organometallic chemistry occupies a central position in organic synthesis because
such systems offer crucial stoichiometric and catalytic routes to the making and breaking
of C–H and C–C bonds.
Ferrocene, bis(cyclopentadienyl)iron, is the most important member of a class of organometallic
compounds known as sandwich compounds. X-ray crystallographic studies on ferrocene have shown
that the iron atom lies equidistant to ten carbon atoms between two parallel planar C5H5
(cyclopentadienyl) rings. Although the original crystallographic result suggested the molecule to be
centrosymmetric, ferrocene and its substituted analogues adopt largely eclipsed structures, with
extremely low barriers to rotation, allowing the rings to spin very rapidly at room temperature even in
the solid state.
C H E M 0 0 2 4 L a b M a n u a l | 17
Each cyclopentadienyl anion, C5H5–, acts as a six-electron donor to the Fe2+ ion. This can be understood
since the ring is made up of five sp2 hybridised carbon atoms each using one electron to bind a
hydrogen atom, two to bind neighbouring carbons, leaving one electron per carbon to donate to the
metal centre, i.e. there are five electrons plus one for the negative charge. Thus, the Fe2+ ion gains
twelve electrons from the cyclopentadienyl ligands in addition to its six valence electrons, so that the
iron achieves the stable 18-electron configuration of krypton.
Ferrocene can be synthesised by initially cracking dicyclopentadiene into cyclopentadiene, C5H6, at
elevated temperatures (a retro-Diels Alder reaction):
Deprotonation of cyclopentadiene using solid KOH forms the cyclopentadienide ion, [C5H5]–, or Cp for
short. The resulting potassium cyclopentadienide is then reacted with iron (II) chloride.
FeCl2.4H2O + 2KC5H5 → [Fe(C5H5)2] + 2KCl + 4H2O
Ferrocene forms thermally stable and volatile orange crystals that can be purified by sublimation or
recrystallisation. In solution, ferrocene is readily oxidised to the blue ferrocenium ion, [Fe(C5H5)2]+. In
some substituted ferrocenes the redox potential of this process has been found to be sensitive to the
glucose concentration in solution and this forms the basis of a glucose sensor used widely by diabetics
in the United States.
One of the key issues in organometallic chemistry is the way in which the chemistry of the ligands is
altered by the presence of the metal. This practical will be concerned with the functionalization of the
Cp by a C-H substitution reaction using a dehydrogenative Heck reaction. Due to synergistic back
bonding between the iron centre and the cyclopentadienyl rings, ferrocene behaves as an electronrich aromatic compound. Ferrocene is 3 x 106 times more reactive than benzene towards electrophilic
substitution. In effect an electrophilic aromatic substitution reaction replaces an ‘H’ atom with another
group, in this case with the electrophile. This reaction can be viewed as a C-H functionalisation
reaction.
18 | C H E M 0 0 2 4 L a b M a n u a l
There are other modern ways of performing C-H functionalisation reactions and in this practical we
use a Pd(II)(OAc)2 catalyst to do this. The complex Pd(II)(OAc)2 is particularly good at C-H
functionalisation of aromatic groups and the reaction generally proceeds by a ‘concerted metalation
deprotonation’ mechanism. The resultant aromatic palladium(II) species can then be used in standard
palladium-catalysed reactions.
In the reaction in this practical the dimethylamino group, coordinatively assists and directs the C-H
functionalisation reaction, making the overall reaction much faster. The resultant organo palladium(II)
species is then used in a Heck reaction. The Heck reaction is stereoselective in most cases for the Ealkene and can also be viewed as a C-H functionalisation reaction with respect to the alkene
component.
Note that normally in a Heck reaction the organopalladium(II) intermediate is accessed by oxidative
addition of Pd(0) into a carbon-halogen or carbon-triflate bond (familiarise yourself with this by looking
up the Heck reaction in your course textbook (Ref 1, 2). You will learn more about this reaction in
CHEM0034. In the reaction in this practical the organopalladium(II) intermediate is accessed from a CH insertion reaction, so does not require the regiospecific preparation of the corresponding halide or
triflate beforehand (familiarise yourself with the mechanism of the reaction in this practical (Ref 4).
Reactions of this sort, that formally involve two C-H functionalisation events to form a new bond, are
sometimes referred to as dehydrogenative coupling reactions. For the coupling reaction to proceed,
two hydrogen atoms are lost, formally as dihydrogen, but usually sequestered as other separate
hydrogen species in the reaction by-products.
C H E M 0 0 2 4 L a b M a n u a l | 19
Objectives
•
•
•
•
To functionalise a Cp ligand.
To use a variety of techniques to purify your compounds.
To assess the success and purity of your reactions by the m.p. and appearance of your product
(by comparison to the literature), and most importantly by using NMR and IR spectroscopy.
To determine the magnetic moment of a transition metal complex by the Evans NMR method.
Pre-Lab (revisit p10-p13 for details)
•
Task 1: Complete your COSHH and risk assessments. For each practical the risk assessment, minus
the chemical hazards, are given to you on Moodle. You are expected to look up the COSHH
assessments for the chemicals that you will be using and fill the chemical hazard and chemical
control sections in. You should then print your risk assessment and stick it in your lab book. You
must also complete the pre-lab safety quiz on Moodle before the start of each practical.
•
Task 2: Provide detailed reaction “equations”, showing all reagents solvents and catalysts.
Calculate the amounts of all reagents in mmol (or in mg where mmol values are given) and the
maximum (100%) theoretical yield based on the limiting reagent.
•
Task 3: Predict how many different chemical environments you would expect to see in the 1H NMR
of ferrocene, dimethylaminomethyl ferrocene and the final Heck product (ignoring R1). Draw the
compounds and label the different environments. For the dimethylaminomethyl ferrocene
starting material, fully assign the 1H NMR spectrum.
•
Example: Ethyl acetate
1
H NMR (300 MHz, CDCl3) δ 1.26 (3H, t, J = 7.0, OCH2CH3), 2.05 (3H, s, C(O)CH3), 4.12 (2H, q, J
= 7.0, OCH2CH3).
Note – J values should be calculated to 1 d.p. Chemical shifts should be quoted to 2 d.p. for 1H NMR
•
Further information on how to assign NMR spectra can be found in Clayden (chapters 13 and 18)
and in the Proton Nuclear Magnetic Resonance Spectroscopy (1H NMR) Guide and Practice on
Moodle (Useful videos and resources).
20 | C H E M 0 0 2 4 L a b M a n u a l
Practical
Important Notes
On day one you should start both parts. Half the class will start with part 1 and half with part 2. See
Moodle lab group allocations for details of which part you should complete first, and which alkene to
use for part 2. Part 2 requires an overnight reaction so it is imperative that you finish part 1 in good
time to set the overnight reaction up (~1 h). For the overnight reaction you will need to complete an
overnight form and get this signed by the senior demonstrator. On day two you should isolate and
purify your product from part 2.
Part 1. Oxidation of ferrocene to ferrocenium hexafluorophosphate
In a round bottom flask equipped with a magnetic stirrer, dissolve 0.50 g of ferrocene in 5 mL of
acetone and slowly add 10 mL of water. Add 0.59 g solid anhydrous FeCl3. Stir the mixture for 15 min.
Vacuum filter the solution through a fritted Hirsch funnel loaded with 1 cm depth of Celite. Add 0.58 g
of NH4PF6, swirl to dissolve, and after 5 min add 5 mL of ethanol. Cool on ice. Filter off the deep blue
product. A second crop may be obtained by reducing the volume of the solution on the rotary
evaporator and adding more ethanol.
The product may be recrystallised if necessary from acetone by addition of ethanol and cooling in an
ice/salt bath.
Record the yield. Record the infrared spectrum (solid). Determine the magnetic moment of the
material by the Evans NMR method (see Appendix 3). You should include details of your calculation in
your lab book.
Part 2. C-H Activation and Heck Reaction
You will have been assigned one of the following alkenes – assignments can be found on Moodle or on
the notice board outside of the Turner lab.
In a 25 mL flask, dissolve N,N-dimethylaminomethyl ferrocene (1.5 mmol) in DMF (3 mL). Add Pd(OAc)2
(5 mol %, 16.8 mg, sign out from stores), K2CO3 (0.45 mmol), tetra-n-butylammonium bromide (0.75
mmol), pivalic acid (1.5 mmol) and your assigned alkene (4.5 mmol). Caution: the alkenes are irritants,
therefore please weigh out in the fume cupboard and keep the flask stoppered at all times. Leave
any pipettes you may have used to dispense the alkene in the fume hood. Complete an overnight
form and get this signed by the senior demonstrator. Stir the mixture at 80 °C overnight. After this time
allow the reaction to cool to room temperature. Then pour the reaction mixture into a separating
funnel, wash out the flask with EtOAc (10 mL) and add this and an additional 20 mL of EtOAc to the
C H E M 0 0 2 4 L a b M a n u a l | 21
separating funnel. Wash with saturated sodium bicarbonate solution (20 mL) (Caution: effervescence)
and then water (2 x 20 mL). Dry the organic layer over MgSO4, filter into a clean, dry, pre-weighed 100
mL flask and remove the solvent on the rotary evaporator. Record your crude yield and crude NMR.
Check the purity of your product by TLC:
(1) Draw (lightly, so as not to disturb the silica!) a pencil line ~1 cm from the bottom of a small silica
gel TLC plate (2.5 x 7.5 cm) and pencil three crosses ~0.5 cm apart, leaving at least 0.5 cm each
side of the outer spots.
(2) In two small specimen tubes, dissolve separately ca. 10 mg of starting material (N,Ndimethylaminomethyl ferrocene) and the final product in dichloromethane (0.5 mL). Using a
capillary tube, spot the starting material solution on to the plate at both the left position and
central position. Similarly, with a fresh capillary tube, spot the reaction solution onto the plate at
the right and central positions.
(3) In a beaker wide enough to house the TLC Plate, add to a depth of less than 1.0 cm the developing
solvent (EtOAc/NEt3 10/1 v/v, Care! Noxious vapour, use in fumehood) and cover with a watch
glass.
(4) Place the plate carefully into the beaker, and develop the plate until the solvent front has travelled
to within 1.0 cm below the top of the plate. Remove the plate from the beaker and mark the
solvent front with a pencil. Visualize the developed plate under UV light, ringing the spots with a
pencil and develop the plate with potassium permanganate solution. Make a sketch of the plate
and calculate the values of the retention factor, Rf, of the compounds. (Measure from the centre
of the spot to the base-line.)
b ef ore
a fter
so lve nt f ro nt
Watch Glass
TLC Plate
Beaker
start
Pencil line
Solvent
Rf =
dist an ce spot
distance
solvent front
22 | C H E M 0 0 2 4 L a b M a n u a l
Purification of your product by Column Chromatography
There are many ways of purifying organic compounds but one of the most
commonly used techniques is column chromatography. Your will use this technique
to purify your crude reaction mixture.
Preparation and elution of the chromatography column should take place in a
fumehood, as flammable solvents are used do not set up or perform in a fumehood
where any reaction involving heating is present. Consult a demonstrator if you are
unsure of what to do.
CAUTION: silica particles cause injury to lungs by inhalation, always handle in a
fumehood. Close the tap at the bottom of the column. Pour silica into the column
to a height of ~15 cm. Carefully transfer the silica in the column to a conical flask
and prepare a slurry by adding your EtOAc/NEt3 10:1 v/v solvent system (Care!
Noxious vapour, do not breathe in, only use in fumehood). Pour the silica slurry into the column using
a powder funnel. Allow the column to stand for a few min and rinse down any silica stuck on the sides
of the column with eluent.
Do not begin the following section with less than 75 min remaining before the end of the lab session
– once you have placed your sample on the column the elution must be completed.
Dilute the reaction product with the minimum amount of eluent so it is mobile. Drain the column until
the level of the solvent is at the top of the stationary phase. Carefully transfer the reaction product
from the round bottom flask to the column using a glass pipette – rinse the flask with ~2 mL of eluent
and transfer the washings to the column. Open the tap and slowly allow the column to drain until the
level of the solvent is at the top of the stationary phase. Add a 1 cm layer of sand on top of the silica
and add your eluent to the column taking care not to disturb the sand and silica as your pour the eluent
in.
Apply some gentle pressure at the top of the column (using compressed air – consult a demonstrator).
NEVER secure the compressed air to the column e.g. with a clip, as the pressure can build up
dangerously. Keep an eye on the column and under no circumstances allow it to run dry! Continue to
elute the column using your solvent system and collect fractions of the eluting solvent. Check each
column fraction by TLC to see which fractions contain your product (use the large, 5 x 10 cm, TLC plates
landscape and spot multiple column fractions on each plate). Visualize the developed plate under UV
light, ringing the spots with a pencil and develop the plate with potassium permanganate solution.
Combine the fractions which contain pure product by TLC into a pre weighed round bottomed flask
(remember that these fractions contain NEt3 so should be kept in a fumehood). Stopper the flask
before taking it to the rotary evaporator to remove the eluent. Check that you have removed all of the
higher boiling NEt3 solvent by weighing the flask and comparing the mass to the maximum theoretical
yield. If your mass is greater than the theoretical yield return the flask to the rotary evaporator.
Record the yield of your product, an IR spectrum, the melting point (if solid) and a 1H NMR, using CDCl3
as solvent. Look up the literature m.p.
C H E M 0 0 2 4 L a b M a n u a l | 23
Results
•
Task 5: Calculate percentage yields for the compounds obtained in parts 1, and 2. For each
compound synthesised attach all spectra in your lab book.
•
Determine the magnetic moment of the material by the Evans NMR method (see Appendix 3).
You should include details of your calculation in your lab book. There is further information on
calculating the magnetic moment on the Moodle page (under useful videos and resources).
•
Task 6: Explain how your data supports the structure of what you should have made. Simply
stating that the product is correct because the data matches the reference spectrum is not
sufficient (See Appendix 4). Comment on any discrepancies (is there still starting material present
or are there impurity peaks such as residual solvent?). In all cases compare your data with those
for authentic material (reference spectra available on Moodle).
References
Keywords: Ferrocene, Organometallic chemistry, Evans magnetic moment, Heck reaction, C-H
activation
(1) Clayden, Greeves, Warren and Wothers, Organic Chemistry 2nd edition, OUP, 2012, ISBN 978-019-927029-3.
(2) Housecroft, P. C. and Sharpe, A. G. Inorganic Chemistry, 4th Edition, Pearson, 2012: Bonding in
ferrocene pp. 887-899 and pp. 924-930, Magnetic moments of transition metals pp. 699-705,
Heck reaction pp. 918-919.
(3) Dimethylaminomethyl ferrocene synthesis: Arthurs, R. A.; Ismail, M.; Prior, C. C.; Oganesyan, V.
S.; Horton, P.; N.; Coles, S. J.; Richards, C. J. Chem. Eur. J. 2016, 22, 3065.
(4) Dehydrogenative Heck reaction with ferrocene: Pi, C.; Li, Y.; Cui, X.; Zhang, H.; Han, Y.; Wu, Y.
Chem. Sci. 2013, 4, 2675.
24 | C H E M 0 0 2 4 L a b M a n u a l
Practical A Mark Scheme (Tasks 5 and 6)
Not Attempted/0
Emerging/1
Developing/2
Satisfactory/3
Exceptional/4
Task 5. Justification of Not
present
or Purity and yield are
Yield and Purity
completely irrelevant. suitable, but yield is
calculated incorrectly.
OR a falsified yield is
OR No product is
provided.
obtained
without
explanation.
Yield is suitable and
calculated
correctly.
However, low purity
(based
on
characterization data).
Yield is
correctly.
Task 6. Evidence of Not
present
or Results compared to
Experimental
completely irrelevant references, but no
Outcome
explanation of how
the data supports the
product structure (or
success
of
the
reaction).
Some
attempt
at
justifying
spectral
differences between
starting material and
product,
and
a
justification of how the
data supports the
product structure (or
success of the reaction)
Good
attempt
at
detailing
spectral
differences and how
that supports the
product structure (or
success
of
the
reaction).
Mostly
correct or comparison
and justification nearly
complete.
calculated Product is obtained in
exceptional yield and purity.
-AND EITHERProduct is obtained in
suitable yield and
purity. -OR- Possible
reasons for the low
yield or purity are
provided.
Comprehensive and correct
comparison of spectral data
to prove the product
structure (or success of the
reaction).
C H E M 0 0 2 4 L a b M a n u a l | 25
PRACTICAL
B:
REACTIONS
OF
CARBOHYDRATES AND COMPLEX NMR
Learning Objective
To chemoselectively synthesise a protected carbohydrate and perform a functionalization reaction
using free radical chemistry. Rationalise the selective protection of carbohydrates and understand how
orthogonal protecting groups can be removed in the presence of others. To rationalise the selectivity
of a free radical functionalization reaction. To understand the different reaction conditions required
for these transformations and also why certain methods/techniques are used to isolate and purify
products. To be able to predict key differences in the NMR spectra of starting materials and the
products and fully assign the spectra of the products using a variety of NMR techniques.
Relates To Course Material
•
•
Carbohydrate chemistry CHEM0034 (TS lecture notes)
NMR CHEM0016 (HCH lecture notes)
Introduction
Carbohydrates
Carbohydrates are often synonymous with energy and metabolism due to the important role of
glucose in powering the Krebs tricarboxylic acid (TCA) cycle and generating high energy biosynthetic
intermediates. However, sugars additionally form the main structural backbone of DNA and RNA and,
whether they are working alone or combined with further biomolecules including lipids and proteins,
mediate several complex cellular processes. Understanding and exploiting the chemistry of
carbohydrates enables new research relevant to carbohydrate-biomolecule interactions which is
essential in developing new antibiotics, antivirals, biologics and anti-cancer agents.
For chemists to prepare specific carbohydrate-based medicines it is essential that we can differentiate
between several hydroxyl groups of similar reactivity, in order to single out specific positions for
modification. Thankfully several methods exist to selectively modify a 1° hydroxyl group in preference
to 2o hydroxyl groups (using bulkier reagents) and several other features, such as the orientation of
one hydroxyl group relative to another (e.g. cis or trans) further aid in discrimination. In addition to
distinguishing between different hydroxyl groups, introduction of hydrophobic protecting groups
improves the solubility of highly polar carbohydrates in common organic solvents.
This practical will be concerned with selective protection and functionalization of glucose. Two glucose
hydroxyl groups are first protected simultaneously through formation of a 4,6-O-benzylidene acetal.
26 | C H E M 0 0 2 4 L a b M a n u a l
Regio-selective cleavage of the benzylidene acetal affords a 6-bromo, 6-deoxy glucose analogue. The
1° halide can be substituted with various nucleophiles into more elaborate structures. In the first step
of the reaction sequence a 4,6-O-benzylidene acetal is introduced.
This reaction employs benzaldehyde dimethyl acetal in the presence of an acid catalyst and proceeds
to form the stable trans decalin-like structure with the phenyl ring occupying the equatorial position
of the newly formed 6-membered ring. This serves to minimise unfavourable 1,3-diaxial interactions.
Next, in what is often referred to as the Hanessian-Hullar reaction, the benzylidene acetal undergoes
regioselective ring opening while converting the 6-OH group to a 1° bromide. The reaction is initiated
by abstraction of the benzylic hydrogen atom by a bromine radical as shown below.
Radical bromination solvents
Radical bromination reactions traditionally use carbon tetrachloride (CCl4) as the solvent as it has no
hydrogen atoms present that can undergo abstraction by intermediate free radical species. However,
CCl4 is highly toxic and is also an ozone-depleting chemical, so its use in large scale chemical procedures
is very restricted. Alternative chlorinated solvents such as tetrachloroethylene and 1,2-dichloroethane
can often be used as substitutes for CCl4, but these solvents are also toxic and are thought to be
carcinogenic. As a consequence of these issues, the identification of alternative solvents for radical
bromination reactions is becoming necessary.
Trifluorotoluene was selected as a suitable alternative solvent for this free radical bromination
experiment as it is less toxic, and the hydrogen atoms in PhCF3 are unlikely to be abstracted by free
radicals.
C H E M 0 0 2 4 L a b M a n u a l | 27
Complex NMR COSY/DEPT
In this experiment we will introduce two powerful NMR techniques, known as DEPT and COSY, which
can aid in the assignment of signals in 13C and 1H NMR spectra. The details of these experiments are
not important here (they will be covered in CHEMM005 next year) but their interpretation is fairly
straightforward.
DEPT (distortionless enhancement by polarisation transfer) is an experiment that produces a 13C
spectrum in which the signals corresponding to CH3 and CH carbons point upwards, the signals
corresponding to CH2 carbons point downwards, and carbons with no hydrogens attached do not
produce a signal at all. So for ethyl acetate (see below), the DEPT spectrum would have three signals:
positive peaks at 14.2 and 21.0 ppm, and a negative peak at 60.5 ppm.
COSY (correlation spectroscopy) is a two-dimensional NMR experiment which can be used to
determine which protons are coupled to each other in a 1H NMR spectrum, and takes the form of a
contour map. A simple schematic COSY spectrum is shown below, for a compound with four protons
labelled A, B, C, D.
The 1H NMR spectrum appears along
the top and left hand side (in a real
spectrum, the signals will not all be
singlets). The peaks which appear in the
COSY spectrum are of two types. Each of
the protons gives a diagonal peak
(shown in solid blue) along the diagonal
line running from bottom left to top
right. In addition, any protons which are
coupled to each other will show crosspeaks (shown in red outline). Reading
down from each of the peaks in the top
spectrum, we can see that A is coupled
to C; B is coupled to D; C is coupled to A
and D; D is coupled to B and C. If the four
protons are on adjacent carbon atoms,
we can therefore conclude that they are
in the order CHA-CHC-CHD-CHB.
28 | C H E M 0 0 2 4 L a b M a n u a l
The usual process of using a COSY spectrum in assigning a 1H NMR spectrum is as follows.
1. Find a signal which can be assigned with confidence (e.g. signal A in this example) and locate the
diagonal peak corresponding to this.
Peak A
Peak A
Diagonal cross peak for A
2. Trace vertically to find a cross-peak showing coupling to another proton, then horizontally back to
the diagonal (in this case, corresponding to proton C). This means that A is coupling to C, so the protons
are adjacent to each other in the molecule CHA-CHC.
Cross peak
C H E M 0 0 2 4 L a b M a n u a l | 29
3. Repeat step 2, searching for different cross-peaks, until all of the coupling network has been
explored. So in the example you can now see that proton C is coupling to both A and D (CHA-CHC-CHD).
Repeating this using the diagonal cross peak of D shows that proton D is coupling to C and B. Therefore
the connectivity of the molecule must be CHA-CHC-CHD-CHB.
Diagonal cross peak for C
Objectives
•
•
•
•
•
•
To chemoselectively protect a polyfunctionalised molecule.
To perform a chemoselective functionalization reaction using free radical chemistry.
To use a variety of techniques to purify your compounds.
To assess the success and purity of your reactions by the m.p. and appearance of your product
(by comparison to the literature), and most importantly by using NMR and IR spectroscopy.
To use 2D NMR to assign your NMR.
To record all coupling constants of your products in the defined style.
Pre-Lab (revisit p10-p13 for details)
•
Task 1: Complete your COSHH and risk assessments. For each practical the risk assessment, minus
the chemical hazards, are given to you on Moodle. You are expected to look up the COSHH
assessments for the chemicals that you will be using and fill the chemical hazard and chemical
control sections in. You should then print your risk assessment and stick it in your lab book. You
must also complete the pre-lab safety quiz on Moodle before the start of each practical.
•
Task 2: Calculate the amounts of all reagents in mmol and the maximum (100%) theoretical yield
based on the limiting reagent.
•
Task 3: Assign the NMR and IR spectra of the starting material (reference spectra are below) in
the following format:
30 | C H E M 0 0 2 4 L a b M a n u a l
Example: Ethyl acetate
IR: 2980 cm-1 (C-H stretch), 1710 cm-1 (C=O stretch)
Note – only key functional group peaks are needed.
1
H NMR (300 MHz, CDCl3) δ 1.26 (3H, t, J = 7.0, OCH2CH3), 2.05 (3H, s, C(O)CH3), 4.12 (2H, q, J
= 7.0, OCH2CH3).
13
C NMR (75 MHz, CDCl3) δ 14.2 (CH2CH3), 21.0 (CH3), 60.5 (CH2CH3), 171.4 (C).
Note – J values should be calculated to 1 d.p. Chemical shifts should be quoted to 2 d.p. for
H NMR and 1 d.p. for C NMR.
Further information on how to assign NMR spectra can be found in Clayden (Chapters 13 and 18).
Reference Spectra
The NMR and IR spectra of methyl-α-D-glucopyranoside are provided below. 1H NMR spectra were run
in DMSO-d6 on a 300 MHz machine (needed for J value calculations), due to overlap you can assign the
multiplicity of peaks between 3.23-3.50 ppm as multiplets. DEPT shows signals corresponding to CH3
and CH carbons pointing upwards, the signals corresponding to CH2 carbons pointing downwards, and
carbons with no hydrogens attached show no signal at all. You do not need to assign the four specific
peaks in the 13C NMR between 70-73 ppm. As these carbon atoms are all in very similar chemical
environments they can be assigned together. You would need to run additional complex NMR
experiments (e.g. HMBC) to confirm which peak corresponded to which carbon.
The anomeric proton (C1-H) has been labelled on the 1H NMR
spectrum – use this as a starting point to work through the COSY and
assign the other peaks in the spectrum. Remember to calculate the J
values from the 1H NMR spectrum, and record the data in the correct
format (see example above).
The protons on C6 are diastereotopic.
C H E M 0 0 2 4 L a b M a n u a l | 31
IR: Methyl-α-D-glucopyranoside
1
H NMR: Methyl-α-D-glucopyranoside
Anomeric proton on C1
32 | C H E M 0 0 2 4 L a b M a n u a l
1
H NMR expansion: Methyl-α-D-glucopyranoside
C H E M 0 0 2 4 L a b M a n u a l | 33
COSY: Methyl-α-D-glucopyranoside (With diagonal cross peak for proton on anomeric C1 indicated)
34 | C H E M 0 0 2 4 L a b M a n u a l
13
C NMR: Methyl-α-D-glucopyranoside
DEPT: Methyl-α-D-glucopyranoside
C H E M 0 0 2 4 L a b M a n u a l | 35
Practical
Important Notes
Do part 1 on day 1 and part 2 on day 2. You can set up the column required on day 2 near the end of
day 1, ready for your next lab session – stop before loading your compound onto silica and stopper the
column. If you leave a column prepared overnight, you will need to complete an orange overnight form
and get this signed by the senior demonstrator.
1. Benzylidene protection of methyl-α-D-glucopyranoside
Dissolve methyl-α-D-glucopyranoside (1.94 g) in dry acetonitrile (20 mL) and attach a reflux condenser.
Flush the apparatus with N2 by adding a Subaseal to the top of the condenser with a N2 balloon and an
exit needle in the Subaseal in the flask and allow to bleed for 2-3 min before removing the exit needle.
Fill another N2 balloon and swap with the depleted one if required. Stir the mixture vigorously.
Add benzaldehyde dimethyl acetal (2.66 mL) and 10-camphor-sulfonic acid (113 mg) and heat the
mixture to reflux for 45 min under N2 (ensure that the reaction mixture is at reflux – i.e. you can see
solvent condensing on the sides of the flask and in the reflux condenser). After this time allow the
reaction mixture to cool to r.t., and neutralize by adding 3 drops of triethylamine. Concentrate the
reaction on the rotary evaporator.
Dissolve the residue in ethyl acetate (75 mL), wash with brine (50 mL), dry over MgSO 4, filter into a
preweighed 250 mL round bottomed flask and concentrate under reduced pressure. Wash the
resulting crystals with diethyl ether and filter to remove excess benzaldehyde dimethyl acetal.
Record the product yield, m.p. IR and 1H NMR spectra. See Appendix 2 for how to make up an NMR
sample.
2. Radical bromination
Weigh the round bottomed flask before beginning. Dissolve methyl-4,6-O-benzylidene-α-Dglucopyranoside (1.36 mmol) in trifluorotoluene (20 mL), attach a reflux condenser and flush the
system with N2 (as in step 1). Add N-bromosuccinimide (1.58 mmol) and barium carbonate (0.58 mmol)
and ensure that the system is under a positive pressure of N2. Heat the reaction to reflux. Monitor the
reaction progress after 1 h by TLC (80% EtOAc/ petroleum ether, using your starting material as a
reference spot) – see instructions on next page.
b ef ore
a fter
so lve nt f ro nt
Watch Glass
TLC Plate
Beaker
start
Pencil line
Solvent
Rf =
dist an ce spot
distance
solvent front
36 | C H E M 0 0 2 4 L a b M a n u a l
(1) Draw (lightly, so as not to disturb the silica!) a pencil line 1.5 cm from the bottom of a small silica
gel TLC plate (2.5 x 7.5 cm) and pencil three crosses 0.5 cm apart, leaving 0.5 cm each side of the
outer spots.
(2) In a small specimen tube, dissolve ca. 10 mg of starting material (methyl-4,6-O-benzylidene-α-Dglucopyranoside). Using a capillary tube, spot the starting material solution on to the plate at both
the left position and central position. Similarly, with a fresh capillary tube, spot the reaction
solution onto the plate at the right and central positions.
(3) In a beaker wide enough to house the TLC Plate, add to a depth of 1.0 cm the developing solvent
(80 % EtOAc/petroleum ether) and cover with a watch glass.
(4) Place the plate carefully into the beaker, and develop the plate until the solvent front has travelled
to 0.5 cm below the top of the plate. Remove the plate from the beaker and mark the solvent
front with a pencil. Visualize the developed plate under UV light, ringing the spots with a pencil
and develop the plate with potassium permanganate solution. Make a sketch of the plate and
calculate the values of the retention factor, Rf, of the compounds. (Measure from the centre of
the spot to the base-line.)
When the reaction is complete (i.e. there is no starting material present by TLC) allow the reaction
mixture to cool to rt and concentrate the reaction mixture under reduced pressure. Record your crude
mass.
Purification of your product by Column Chromatography
There are many ways of purifying organic compounds but one of the most commonly used techniques
is column chromatography. You will use this technique to purify your crude reaction mixture.
Preparation and elution of the chromatography column should take place in a fumehood, as flammable
solvents are used do not set up or perform in a fumehood where any reaction involving heating is
present. Consult a demonstrator if you are unsure of what to do.
CAUTION: silica particles cause injury to lungs by inhalation, always handle in a
fumehood. Close the tap at the bottom of the column. Pour silica into the column
to a height of ~15 cm. Carefully transfer the silica in the column to a conical flask
and prepare a slurry by adding your 80% EtOAc/petroleum Ether solvent system.
Pour the silica slurry into the column using a powder funnel. Allow the column to
stand for a few min and rinse down any silica stuck on the sides of the column with
eluent.
Do not begin the following section with less than 75 min remaining before the
end of the lab session – once you have placed your sample on the column the
elution must be completed.
Dilute the reaction product with the minimum amount of dichloromethane so it is mobile (~2 mL).
Drain the column until the level of the solvent is at the top of the stationary phase. Carefully transfer
the reaction product from the round bottom flask to the column using a glass pipette – rinse the flask
with ~2 mL of eluent and transfer the washings to the column. Open the tap and slowly allow the
column to drain until the level of the solvent is at the top of the stationary phase. Add a 1 cm layer of
sand on top of the silica and add your eluent to the column taking care not to disturb the sand and
silica as your pour the eluent in.
C H E M 0 0 2 4 L a b M a n u a l | 37
Apply some gentle pressure at the top of the column (using compressed air – consult a demonstrator).
NEVER secure the compressed air to the column e.g. with a clip, as the pressure can build up
dangerously. Keep an eye on the column and under no circumstances allow it to run dry! Elute the
column using your solvent system and collect fractions of the eluting solvent. (Roughly two volumes
of solvent equal to the volume of silica should be collected and then change to 100% EtOAc, again two
volumes of solvent equal to the volume of silica should be collected. See a demonstrator if you are
unsure about this).
Check each column fraction by TLC to see which fractions contain your product (use the large, 5 x 10
cm, TLC plates landscape and spot multiple column fractions on each plate). Visualize the developed
plate under UV light, ringing the spots with a pencil and develop the plate with potassium
permanganate solution. Combine the fractions which contain pure product by TLC into a pre weighed
round bottomed flask and remove the eluent using a rotary evaporator.
Record the product yield, m.p., IR, mass spec and 1H NMR spectra.
Results
•
Task 5: Calculate percentage yields for the compounds obtained in parts 1, and 2. For each
compound synthesised attach all spectra in your lab book. Include m.p. with literature value
comparison and reference if the compound is solid.
•
Task 6: Explain how your data supports the structure of what you should have made (See
Appendix 4). Comment on any discrepancies (is there still starting material present or are there
impurity peaks such as residual solvent?). In all cases compare your data with those for authentic
material (reference spectra available on Moodle). You are also provided with a COSY spectrum of
authentic material, which you should print out. Use this to assist in the assignment of the peaks
and attach your copy of the COSY showing which cross-peaks correspond to which couplings.
References
Keywords: Carbohydrate, protecting groups, COSY Spectra, radical bromination
(1) Clayden, Greeves, Warren and Wothers, Organic Chemistry 2nd edition, OUP, 2012, ISBN 978-019-927029-3.
(2) Crich, D., et al. (2006). “On the regioselectivity of the Hanessian–Hullar reaction in 4,6-Obenzylidene protected galactopyranosides.” Carbohydrate Res. 341(10): 1748-1752.
(3) Radical Bromination: Sato, K.; Yoshimura, J. Carb. Res. 1982, 103, 221.
38 | C H E M 0 0 2 4 L a b M a n u a l
Practical B Mark Scheme (Tasks 5 and 6)
Not Attempted/0
Emerging/1
Developing/2
Satisfactory/3
Exceptional/4
Task 5. Justification of Not
present
or Purity and yield are
Yield and Purity
completely irrelevant. suitable, but yield is
calculated incorrectly.
OR a falsified yield is
OR No product is
provided.
obtained
without
explanation.
Yield is suitable and
calculated
correctly.
However, low purity
(based
on
characterization data).
Yield is calculated Product is obtained in
correctly.
-AND exceptional yield and purity.
EITHER- Product is
obtained in suitable
yield and purity. -ORPossible reasons for
the low yield or purity
are provided.
Task 6. Evidence of Not
present
or Results compared to
Experimental
completely irrelevant references, but no
Outcome
explanation of how
the data supports the
product structure (or
success
of
the
reaction).
Some
attempt
at
justifying
spectral
differences between
starting material and
product,
and
a
justification of how the
data supports the
product structure (or
success of the reaction)
Good
attempt
at
detailing
spectral
differences and how
that supports the
product structure (or
success
of
the
reaction).
Mostly
correct or comparison
and justification nearly
complete.
Comprehensive and correct
comparison of spectral data
to prove the product
structure (or success of the
reaction).
C H E M 0 0 2 4 L a b M a n u a l | 39
PRACTICAL C: SYNTHESIS OF CRYSTALLINE
FUNCTIONAL MATERIALS
Learning Objective
To synthesize crystalline functional materials and to apply diffraction techniques for phase
identification of the reaction products. To understand the different reaction conditions required for
the synthesis of main group and transition metal bearing materials. To learn modern techniques
applied in the synthesis of crystalline inorganic compounds and techniques required for handling air
sensitive materials. To interpret X-ray diffraction patterns and infrared spectra. The lithium iron
phosphate (LFP) materials synthesised in this experiment are similar to those you used in CHEM0025
to construct a fully operational rechargeable Li-ion battery.
Relates to Course Material
•
•
•
Crystallography and X-ray diffraction in CHEM0013 (BS lecture notes) and CHEM0014 (JKC lecture
notes).
Ion intercalation and conduction in CHEM0039 (YX lecture notes).
Electrochemistry and battery materials (CHEM0025).
Introduction
In this experiment you will be working on the synthesis and characterisation of crystalline inorganic
functional solids. These ionic materials perform a predefined “function”; for instance, the magnetic
moment of ferromagnetic oxides below the Curie temperature can be oriented in two (up and down)
directions and used to represent the two states (0 and 1) of a bit in computer memories.
The practical work is based on two phosphate materials: hydroxyapatite (HA) Ca5(PO4)3(OH) and
lithium iron phosphate (LFP) LiFePO4. HA is a biomaterial that mimics the structure and composition
of bones, and LPF is the cathode material currently used in commercial rechargeable Li-ion batteries.
LFP gives reversible electrochemical intercalation of Li-ions, which can be exploited in electrochemical
cells for energy storage.
Both materials have relatively complex crystalline structures (shown in Figure C.1), enabled by the
chemical flexibility of the molecular phosphate ions (PO43-) bridged by octahedral units of the cations
(Li, Ca, and Fe). Despite the chemical similarity due to the phosphate anion, the presence of the redox
active Fe-ion in LFP requires a strict control of the synthesis conditions that you will be required to
understand and apply.
The structural flexibility of HA and LFP also enables chemical modifications of both materials by doping,
which can be applied to control and improve the functional behaviour. The description provided in the
pages below relates to the synthesis of undoped HA and LFP. Students who continue with the research
project in Expt. C2 will be asked to define and attempt the synthesis of doped LFP. Because of the
complexity of solid-state synthesis, not all synthesis attempts will be successful in yielding the target
HA and LFP compounds in a pure form; this is a common situation in materials research. Understanding
40 | C H E M 0 0 2 4 L a b M a n u a l
if the synthesis has been successful and why it has (not) been successful is an important learning
objective. Crystalline solids are studied primarily with diffraction techniques, and you will learn how to
prepare your samples for powder X-ray diffraction (PXRD) studies and how to analyse the resulting
diffraction patterns to identify the phases produced.
Figure C.1. Crystal structures of HA (top) and LFP (bottom) highlighting the bridging role of the
phosphate ions PO43- in connecting MO6 octahedra of the cations (M = Ca, Fe, and Li).
Hydroxyapatite
Bioceramics are inorganic materials specifically formulated for reconstruction of damaged sections of
the skeletal system due to their ability to form a stable interface with surrounding tissues and preform
efficiently the natural tissue. In general, bioceramics can be described according to the tissue response
as bioinert, bioactive and/or bioresorbable, as detailed in Table 1.
Table 1. Bioinert, bioactive and bioresorbable materials.
Bioceramic type
Properties
Example
Bioinert
Tissues form a non-adherent
fibrous capsule around the
implant
Alumina (Al2O3), Zirconia (ZrO2)
and carbon
Bioactive
Tissues form an interfacial
bond with the implant
Hydroxyapatite, Bioglass
Tissues replace implant
β-Tricalcium phosphate,
carbonated hydroxyapatite,
calcium carbonate
Bioresorbable
C H E M 0 0 2 4 L a b M a n u a l | 41
Calcium phosphates are the most widely used bioceramics for implants mainly due to their
resemblance to bone mineral, and their ability to form strong chemical, physical and biological bonds
with the tissue interface. They are well known for their use as bone graft substitutes, coatings on
metallic implants, gene therapy, reinforcements in biomedical composites and bone, and as
components in dental cements.
HA and tricalcium phosphate (TCP) are amongst the most common calcium phosphates. Synthetic HA
is bioactive and similar to biological apatite, the main mineral constituent of teeth and bone. HA is
biocompatible, bioactive and has a low solubility in wet media. This bioceramic has been employed as
a scaffold material to encourage new bone formation for osteoinductive coatings on metal implants
and as a bulking agent for bone fillers. In contrast, beta-tricalcium phosphate, β-TCP, [Ca3(PO4)2],
another bioceramic, has a higher dissolution rate compared to HA in vivo. A range of calcium
phosphates can be obtained with different Ca/P molar ratio and pH stability ranges (see Table 2).
Multiple techniques have been used for the preparation of HA including solid-state synthesis, coprecipitation method, sol-gel technique, emulsion synthesis, and batch hydrothermal synthesis.
Table 2 – Calcium phosphates (from Dorozhkin, 2010).
Lithium Iron Phosphate
The structure of LFP is shown earlier in Figure C.1. The FeO6 and PO4 polyhedra connect to form
channels oriented in the (001) crystallographic direction, where Li-ions are located. When the LFP
sample is connected to an electric circuit, applying a voltage can cause Li-ions to migrate reversibly
inside and outside the material. The process is called intercalation and is at the basis of energy storage
in Li-ion batteries. Much more detail about the electrochemistry of Li-ion batteries and the material
chemistry of LFP that enables this application will be discussed in the instrumental work of CHEM0025,
where the LFP materials synthesised here will be employed to build a fully operational rechargeable
battery. The same process will also be investigated via computer simulation in the solid-state computer
modelling experiment (Instrumental 2 part of CHEM0025). An overview of Li-ion batteries and the
materials employed can be found in Goodenough, 2013.
Traditionally, inorganic solid materials have been obtained by mixing binary compounds of the
individual cations in the required stoichiometric ratio, grinding the mixture, and calcining at high
42 | C H E M 0 0 2 4 L a b M a n u a l
temperature (1000˚C or above to enable interdiffusion of the cations) for a long period of time. Often
the procedure must be repeated several times to achieve homogeneous mixing of the components in
a ternary or more complex solid. The length of such an experimental procedure is unsuitable for
undergraduate classes. However, recent research has shown that solids with even complex
compositions can be obtained efficiently with solution-based methods that employ non-ambient
conditions. You will employ two such methods in the experiment: the first is hydrothermal synthesis,
in which a water solution with stoichiometric composition is heated in an autoclave (a small pressure
cooker), reaching temperature and pressure well above the normal boiling point of water. The second
method is based on microwave irradiation, in which energy is transferred to the sample very rapidly in
a microwave oven, enabling efficient mixing and phase transformation. The products of the autoclave
and microwave synthesis will often need overnight heating and calcination processes to ensure that a
phase with a high degree of crystallinity is obtained. Learning how to select the correct conditions and
how to operate these modern synthetic tools will be another important learning objective of the
experiment.
Objectives
•
•
•
•
To apply microwave and hydrothermal methods in the synthesis of functional phosphate materials
To apply and interpret diffraction techniques for phase identification of the solid products.
To use infrared (IR) spectra for the identification of functional groups in phosphate materials
To understand the different reaction conditions required for the synthesis of main group and
transition metal bearing materials.
Pre-Lab Tasks
Task 1. Safety Assessment
•
•
Complete your COSHH and risk assessments.
Complete the pre-lab safety quiz on Moodle.
Task 2. Table of Reagents
•
•
•
•
Write the formula and calculate the mass of calcium nitrate tetrahydrate and ammonium
phosphate dibasic required to make 50 mL of solutions 0.5000 M in Ca and 0.3000 M in PO 43-.
These solutions will be employed for the HA synthesis.
Calculate the mass of iron sulphate heptahydrate and of orthophosphoric acid solution needed to
make 25 mL of solution 0.5000 M in Fe and PO43-. Similarly, calculate the amount of lithium
hydroxide monohydrate required to make 25 mL of solution 1.500 M in Li. These solutions will be
employed for the microwave synthesis of LFP.
Orthophosphoric acid is dispensed in the laboratory as 85.0 weight percent solution.
Write balanced equations for the synthesis of HA and LFP using the precursors listed above.
Task 3. Pre-Lab Questions
1. Why is orthophosphoric acid measured by mass and not by volume since it is dispensed as
solution?
2. Why is important to have reflux setup in the microwave synthesis?
3. Why is LFP dried in a vacuum oven, but HA is not?
C H E M 0 0 2 4 L a b M a n u a l | 43
4. What important information can be extracted from the X-ray diffraction patterns of the samples?
Representative XRD patterns of HA before and after high-temperature calcination are shown
below. What happens during calcination and why are the peaks sharper in the calcined sample?
Figure C.2. XRD patterns of HA after hydrothermal synthesis (as prepared) and calcination treatment
at 1000˚C for 1 h (heat treated) compared with the reference pattern in structure databases (bottom).
Task 4. Lab Notebook
•
This task will be completed alongside your two-day experiments. Make sure it is signed off by a
demonstrator before the end of Day 2 session.
Practical
Schedule
This practical contains overnight synthesis and requires you to use equipment at specified times.
Consequently, you must ensure you follow the schedule below to successfully complete the lab; there
is no room in the schedule for any additional overnight experiments. You will work individually and
must perform the XRD sample preparation on the specified day, and the appropriate diffractometers
will not be available at other times.
Day 1: Materials synthesis: you will perform the microwave and hydrothermal syntheses of HA and
the microwave synthesis of LFP. By the end of Day 1, you must: (i) have left the hydrothermal
synthesis of HA in the overnight oven; (ii) have submitted the product of the microwave
synthesis of HA for calcination; (iii) have performed the IR measurement on the product of the
44 | C H E M 0 0 2 4 L a b M a n u a l
microwave synthesis of HA before calcination; (iv) have left the product of the microwave
synthesis of LFP in the vacuum drying oven.
Day 2: Materials recovery and characterisation: you will recover the product of the hydrothermal
synthesis of HA and leave it in the drying oven. You will perform a series characterisation on the
HA and LFP products. By the end of Day 2, you must: (i) have performed the IR measurement
on the calcinated HA product, the dried hydrothermal HA product, and the overnight dried
microwave LFP product; (ii) have prepared PXRD samples of the calcinated HA product and the
overnight dried microwave LFP product.
Important Notes
General: All preparations should be performed in a fumehood (in particular, ammonium hydroxide
should not be removed from the fumehood). Instructional videos for several of the procedures used
in this lab (microwave use, autoclave use, and XRD sample preparation) are available on Moodle under
the tab “Useful Videos and Resources”. You should watch these videos before you start the lab.
Timings: To successfully complete this lab, which involves an overnight step, you must ensure that you
complete the correct tasks at the correct time. You may also only use the diffractometers at the
specified times. These are all detailed in the schedule section.
Microwaves: The water flow to the microwave condensers has been pre-set by the technical staff and
should not be adjusted; if there is a problem with the water flow, please consult a demonstrator or
technician. Do not put cork rings or quick-fit clips in the microwave. When preparing your microwave
reactions, do not grease your glassware, do not add anti-bumping granules to your reactions, and
ensure you have removed the magnetic stirring bar.
Autoclaves: When you sign out an autoclave from stores, you will be required to confirm that you have
watched the video of autoclave use on Moodle. If you have not watched it, you must ask a
demonstrator to go through it with you before you are given an autoclave. A demonstrator must be
present when you assemble the autoclave before placing it in the oven.
Ovens: All ovens have sample sign in/out sheets on them. Please use these so that we know what is
in the ovens and can keep track of samples. To place/remove samples in the vacuum oven, ask a
demonstrator for assistance.
XRD: you must watch the videos of PXRD sample prep…