Understanding Microwave Sample Preparation for USP Methods 232,233 and 2232
USP Method 231
Current Methodology
• USP Method 231 – introduced in 1905
– 3 different sample preparation methods can be used
dependent on sample type
– Metal impurities: Ag, As, Bi, Cd, Cu, Mo, Hg, Pb, Sb,
Sn are precipitated with sulfide and compared to lead
standards
• Qualitative (semi-quantitative at best) test to
determine metal impurities
Limitations of USP Method 231
Methodology
• Difficult to reproduce results
– Solutions and standards change with time
• Nondiscriminatory test – Individual metals not reported separately, reported as group
• Low spike recoveries – Aggressive sample preparation techniques result in low
recovery of volatile metals
• Large sample size of 2 grams is required to
achieve the MDL of 10 ppm
New USP Chapters Will Replace Method 231
In May 2014
• Chapter 232 - Elements of Interest and Limits
• Chapter 233 – Procedures
• Chapter 2232 – Dietary Supplements Speciation*
* Implementation of 2232 may be delayed
USP 232 Elements of Interest
• The Big Four Toxic Elements:
– Always must be analyzed – As, Cd, Pb, Hg
• Environmental and Catalyst Impurities:
– Must be analyzed if added during the process either
intentionally or unintentionally
– Cr, Cu, Mn, Mb, Ni, Pd, Pt,V, Os, Rh, Ru, In
USP 233 Sample Preparation Options
• Sample preparation procedures include:
– Direct aqueous solution – if the sample dissolves in DI water,
then acidify samples per instrument manufacturer specifications
and perform analytical measurement
– Direct organic solution – if the samples dissolves in organic
solvent, perform analytical measurement (if instrument is
organic solvent compatible)
– Indirect solution (closed vessel digestion) – if sample will not
dissolve in DI water or organic solvent, then closed vessel
digestion is required
USP Method 233
Sample Preparation – Closed Vessel Digestion
• Choice of strong acid (HNO3, H2SO4, HCl, HF, Aqua
Regia) is dependent on the sample matrix
• Sample weights and acid volumes can be adjusted for the
sample type and digestion apparatus
• Closed vessel digestion procedure parameters to be
determined by user
Why Do We Digest Samples?
• Measurements take time
• Signal must stay constant during measurement
• Analyzer needs homogeneity
• The solution is a solution…..
Why Pressurized Digestion?
• Temperatures above boiling point
• Oxidative potential of reagents is higher at
elevated temperatures
• Digestion is faster and more complete
Why the USP Pursued a
Microwave Method?
• Rapid Heat Up Time
• Instantaneous on:off = control
• Reproducibility of conditions
• Only the sample and reagent is heated
• Rapid cool down
• High Throughput
CEM’s Solutions to Sample Preparation for
USP Method 233
• Applications Support
– Free phone support for applications issues and method development
– Applications notes, reference papers and sample preparation advice available
24-7 on the CEM website.
– ICP-OES and ICP-MS capabilities for analytical measurement
– IQ/OQ/PQ Documentation
– CEM has been attending USP meetings regarding method 232, 233 for three
years in order to provide our customers the best and latest advice regarding
the changes being made
CEM IQ/OQ/PQ and Service Capabilities • CEM service offers IQ/OQ/PQ for SP–D and Mars
• CEM service offers service contracts for SP–D and Mars that
include recertification of the instrument Discover SP-D
Microwave Accelerated Reaction System
IQ, OQ and PQ
Instrument Qualification
Operation Qualification
Performance Qualification
Installation Procedures
CEM Corporation
P.O. Box 200 3100 Smith Farm Road
Matthews, NC 28106-0200 704.821.7015 ▪ 800.726.3331
www.cem.com [email protected]
600099S Rev. 1
CEM Solutions for
USP Method 233- Documentation
CEM Synergy–D Software
Operating Procedures Page 1 – Instrument and Software Start-up Page 3 – Set-up New User Accounts Page 4 – Copy Applications (CEM) Methods to a User Page 4 – Create New Methods Page 6 – Set-up and Start a Sample Queue with the Explorer Page 7 – Set-up and Start a Sample Queue without the Explorer Page 8 – Add a Priority Sample to a Sample Queue Page 9 – Save Explorer Rack Sequences Page 9 – View Data and Generate Reports Page 12 – Automatic Save of Data After Each Run Page 13 – Save Data to a File Instrument and Software Start-up
1. Turn on the Discover SP–D.
2. Double click on the SynergyD icon on your computer desktop.
3. The software will load and a CEM Discover Logon screen will prompt you to select the User and type the
Password.
4. The software will open. Note the difference between the screen with and without the Explorer autosampler.
Make sure the Discover (and Explorer) are connected– see highlighted area.
CEM SP–D Maintenance Procedures Page 1 – Instrument Adjustment – Explorer Attenuator Set-up Page 4 – Instrument Adjustment – Explorer Rack Set-up Page 7 – Instrument Adjustment – Load/Return Vessels, Adjust Vessel Pickup Height, Home Explorer Page 9 – Instrument Repair – Vessel Failure Clean-up Page 13 – Instrument Test – IR Temperature Verification with Water Page 14 – Instrument Maintenance – Spray Grippers and Rails with Teflon Page 15 – Instrument Maintenance – Grease ActiVent Drive Gear Page 17 – Instrument Maintenance – Checking the ActiVent for Corrosion Instrument Adjustment – Explorer Attenuator Set-up
1. Install 35 mL attenuator
2. In CEM SynergyD software, go to top menu bar → Explorer → Explorer Attenuator Calibration. The Explorer Attenuator Setup box will open.
3. Click Calibrate Attenuator Position – see highlighted area.
CEM’s Solutions to Sample Preparation for
USP Method 233
• Customized Training Available on Site or at CEM
CEM’s Sample Preparation
Instrumentation Line-up
CEM SP-D with Explorer
(Sequential Sample Preparation)
CEM Mars6
(Batch Sample Preparation)
SP–D Pharmaceutical Applications
• CEM has created nearly 100
pharmaceutical application notes
so far – constantly adding more
• Samples include raw materials,
stabilizers, excipients, fillers and
finished products
CEM’s Sample Preparation Considerations
For USP 233
• Nitric acid alone will not work for all sample types, even at high
temperatures
• Nitric acid uses: – ~ 200oC will digest most organic samples
– ~ 210oC - 220oC will work for samples with more difficult matrices
• Acid mixtures will be required for most inorganic samples – HCl required for recovery of most metal catalysts as well as Sn and Sb
– HF needed for silicates – silicates can be found in tablet coatings, fillers, etc.
– Boric acid – dissolve insoluble fluoride due to HF and neutralize HF
• Sample size & Elements of Interest – Will dictate the requirements of ICP-OES or ICP-MS for your laboratory
CEM’s Sample Preparation Considerations
For USP 233
However…
A mixture of 9 ml of HNO3 and 1 ml of HCl will digest and stabilize the
majority of samples and elements for analysis that don’t require HF
Examples of Samples Digested in Nitric Acid and HCl
Antioxidant Tablets Ascorbic Acid Aspirin Beta Carotene Calcium Carbonate
Calpan Chromium Chelate Claritin Cold/Flu Relief Cupric Sulfate
d-Biotin Folic Acid Herbal Supplement Hydrocodone Kelp
Magnesium Oxide Manganese Carbonate Niacinamide Non-Drowsy Allergy Relief Potassium Chelate
Protopic Ointment Pyridoxine Riboflavin Selenium Chelate Thiamine
Valsartan Vitamin B12
Vitamin D3 Vitamin E Zinc Sulfate
Sample Weight: 0.1 - 0.3g
Reagents: 9 ml HNO3 and 1 ml HCl
Sample Preparation Procedure
Microwave Heating Program
Control Style: Ramp To Temperature Pressure Mode: Organic
Stage Temp(C) Ramp
Time
Hold
Time
Pressure
(PSI)
Power (W) Stirring
1 200 5:00 3:00 400 300 Med
Samples Digested in Nitric Acid and HF
Allergy Relief Dicloflenac Potassium Glipizid/Metformin HCL Losartan Potassium Multivitamin
Naproxen Sodium Sleep Aid Sodium Aluminum Silicate Talc
Vitamin C
Sample Weight: 0.1 - 0.3g
Reagents: 9 ml HNO3 , 1 ml HF
Sample Preparation Procedure
Microwave Heating Program
Control Style: Ramp To Temperature Pressure Mode: Organic
Stage Temp(C) Ramp
Time
Hold
Time
Pressure
(PSI)
Power (W) Stirring
1 200 5:00 3:00 400 300 Med
SP–D Blank and Spike Recovery Studies
• Blank and cleaning
studies – extensive
testing at ppb/ppt
levels with ICP-MS
underway
• Recovery studies –
ppm studies
completed and
ppb/ppt studies
underway
• Results available
during next webinar
Multi-Vitamin
(250 mg) Ca Fe Mg Zn Cu Mn
#1 489.3 70.67 181.50 74.500 7.285 7.754
#2 493.2 70.47 175.9 76.31 7.664 8.024
#3 458.1 68.09 178.2 71.74 7.07 7.593
#4 471.7 64.04 182.6 72.41 7.05 7.386
#5 482.6 64.99 173.4 73.35 7.039 7.351
AVG (ppm) 478.98 67.652 178.32 73.662 7.2216 7.6216
True Value
(ppm) 370 66.2 184 55.8 7.360 7.360
% Recovery 128% 100% 96% 131% 96% 101%
Std Dev 14.24 3.06 3.82 1.808 0.267 0.278
%RSD 2.97 4.52 2.14 2.45 3.70 3.65
SP-D Spike Recovery Results of 125 ppb Metal Impurities As Hg Cd Pb Cr Mn Ni Pd Rh V
Acetylsalicylic Acid
Tablet Average 127.5 114.7 118.7 121.3 123.2 120.5 121.5 107.0 132.2 123.8 % Recovery 102 92 95 97 99 96 97 86 106 99 RSD 7.45 1.04 1.21 1.44 0.80 2.62 4.82 3.25 0.81 1.01
Loratadine
Pseudoephedrine
Tablet
Average 90.5 98.2 103.3 98.8 104.2 103.8 99.8 93.7 108.5 100.8 % Recovery 72 79 83 79 83 83 80 75 87 81 RSD 5.88 1.23 1.72 6.72 0.50 3.71 3.16 1.12 5.63 0.29
USP Method 233 Proposed accuracy is 70-150% with
precision < 20%
SP-D Spike Recovery Results of 125 ppb Metal Impurities As Hg Cd Pb Cr Mn Ni Pd Rh V
Folic Acid Average 114.8 118.3 119.3 120.2 122.3 120.7 120.0 119.8 128.3 123.8 % Recovery 92 95 95 96 98 97 96 96 103 99 RSD 4.96 0.86 1.05 5.57 1.76 1.93 1.79 2.45 2.85 0.84
Zinc Sulfate Average 124.7 116.5 121.3 125.3 126.0 113.3 121.5 123.7 136.7 126.3 % Recovery 100 93 97 100 101 91 97 99 109 101 RSD 3.50 1.27 1.61 3.27 3.24 7.02 1.21 1.55 2.13 1.15
USP Method 233 Proposed accuracy is 70-150% with
precision < 20%
Conclusions – CEM SP–D • Fast sample digestions
• Digest real sample sizes – ICP-OES and ICP-MS have plenty of
sensitivity
• Digest a mix of sample types with different acids and temperatures,
ALL during a single run
• Automated, unattended operation
• Several safety features
– Temperature and pressure control
– ActiVent pressure management
– Safety interlocks
• IQ/OQ/PQ and service contracts available
• USP has already purchased the SP-D and Explorer