Testing Pharmaceutical Samples Using Microwave Digestion in Line with USP Chapter <233>

New methods of pharmaceutical sample preparation and analysis have been developed to overcome the limitations of USP Chapter <231> – Heavy Metals published in 1905. The substitutes such as USP Chapters <232> and <233> were proposed in January 2010 and subsequently updated in 2011, 2012, and 2013. The method is expected to be implemented in August 2015 and to come into use in December 2015 during which the USP Chapter <231> will be completely eliminated.

Discover® SP-D

USP Chapter <232> contains fifteen elements, which need to be examined at different concentration limits set depending on the mode of introduction to the body. In general, the “big four” elements – lead, cadmium, mercury, and arsenic are the key elements to be considered owing to their pervasiveness and toxicity.

The excipients, raw materials, and contaminants need to be tested during the manufacture of leachable metallic impurities, catalyst metals, and API’s. Further, depending on the environment, catalysts, and manufacturing process, additional elements are required to be tested.

This article describes the sample preparation and analyte recovery of three pharmaceutical samples added with Hg, Cd, and As.

USP Chapter <232> - Limits of Elemental Impurities

USP Chapter <232> describes the standard range limits of using heavy metals in drug products and active pharmaceutical ingredients with respect to three different administration modes such as inhalation, parenteral, and oral. These elements may be intentionally added during manufacture, transferred through the processing equipment, and naturally present in the components.

USP Chapter <233> - Elemental Impurities – Procedures

The sample preparation and analytical methods of determining heavy metals in finished products and pharmaceutical raw materials are discussed in the USP Chapter <233>. The chapter provides an outline of four sample preparation methods such as Indirect Solution, Direct Organic Solution, Direct Aqueous Solution, and Neat.

The Indirect Solution technique is employed in most of the finished products and pharmaceutical raw materials – which involves dissolving the sample in a closed vessel containing a concentrated acid solution for analysis through an inductively coupled plasma mass spectrometer (ICP-MS) or an inductively coupled plasma optical emission spectrometer (ICP-OES).

When compared to several digestion techniques such as hot blocks and hot plates, microwave digestion instruments offer efficient control of volatile elements, lower acid consumption, and complete destruction of background organics.

In addition, the availability of pressure and temperature control option in microwave digestion systems ensure rapid turnaround of digested samples as well as more reproducible digestion conditions. It also allows appropriate documentation of the preparation procedures of each sample.

CEM provides two different types of microwave systems used for acid digestion: the MARS 6™ that offers high-throughput batch processing of samples and the Discover® SP-D offering an automated, sequential platform. Both the models operate in compliance with 21 CFR Part 11.


Discover® SP-D

The first pharmaceutical sample set was prepared using the Discover® SP-D as it offers excellent flexibility to operate a wide range of samples with automation. In the addition, the presence of an autosampler ensures sequential preparation of each sample independently for analysis purposes, without requiring lab personnel.

The samples can be prepared and cooled within 10 mins using the CEM’s patented Focused™ Microwave technology. A 35mL Pyrex® vessel with a snap-on cap is used for the convenient preparation of pharmaceutical samples. This system is compact with temperature and pressure control options and offers documentation of each sample.


The second sample preparation was carried out using the MARS 6™ integrated with One Touch Technology®. The MARS 6™ consists of sensors at the cavity bottom for identifying the type and number of vessels. This system calculates the optimum power conditions required for ensuring complete digestions and achieving the desired temperature.

Pre-programmed methods enable the user to select the type of sample, and the analysis is carried out independently by the instrument. The system facilitates large throughput batch analysis of up to 40 samples.


The MARS 6™ includes the high- temperature, high-pressure Easy Prep™ Plus vessels in which the samples are digested. DuoTemp™ feature is integrated with a fiber optic probe and Contactless All-Vessel IR Temperature Sensors for efficient monitoring and control of temperature.

This feature automatically chooses the control vessel depending on the reaction condition and performs adjustments during the operation so as to control the temperature with respect to the reactive vessel, yielding reproducible, safe digestions without generating exothermic reactions.

Procedure for Analyzing Samples

Three product samples including a liquid gel capsule with diphenhydramine HCl, a tablet containing acetylsalicylic acid, and another tablet containing loratadine were analyzed during the experiment. Each of the samples is placed in Discover SP-D 35mL vessel or an EasyPrep Plus vessel to which 9mL of concentrated ultra-pure nitric acid and 1mL of a spike solution of deionized (DI) water with 50ppb of cadmium, mercury, and arsenic were added.

In addition, 1mL of hydrogen peroxide was added to the Easy Prep vessel, and 0.5mL was added to SP-D vessel. Table 1 shows the samples and blanks prepared in triplicate using the digestion parameters.

Table 1. Digestion Parameters for MARS 6 and Discover SP-D

  Sample Type Ramp Time (minutes) Hold Time (minutes) Digestion Temperature (°C)
MARS 6 with EasyPrep Plus vessel Organic 15 15 210
Discover SP-D with 35-mL vessel Organic 5 3 210

Samples were then cooled to ambient temperature conditions, diluted to 50mL using DI water and transferred to autosampler vials. Using 20% nitric acid, calibration standards containing cadmium, mercury and arsenic were prepared at concentrations of 0.1, 0.5, 1, 5, 10ppb. The samples were then allowed to run on the Thermo Scientific iCAP Q ICP-MS based on the conditions given in Table 2.

Table 2. Conditions for running sample on the Thermo Scientific iCAP Q ICP- MS

Forward Power 1500 W
Nebulizer Gas 1 L/min
Auxiliary Gas 0.8 L/min
Cool Gas Flow 14.0 L/min
Collision Cell Gas He at 4.5 L/min
Sample Uptake/Wash Time 45 seconds each
Dwell Times Optimized per analyte
Number of Points Per Peak 1
Replicates 3
Internal Standard 1.00 ppm yttrium

Results and Discussion

Table 3. shows the results of the MARS 6™ spike recovery study of 50ppb of pharmaceutical finished products, while Table 4 shows the results of the Discover® SP-D spike recovery study of pharmaceutical finished products.

Table 3. MARS 6™ Spike Recovery Results (ppb) of 50.0 ppb As, Cd, and Hg

  As Hg Cd
Acetylsalicylic Acid Finished Product
Analysis 1 49.04 60.93 48.63
Analysis 2 58.79 46.87 46.21
Analysis 3 57.71 62.29 49.45
Average 55.18 56.70 48.10
Percent Recovery 110.36 113.40 96.19
RSD 9.68 15.05 3.50
Loratadine Finished Product
Analysis 1 73.22 45.93 51.37
Analysis 2 67.40 50.02 58.88
Analysis 3 63.44 58.04 56.75
Average 68.02 51.33 55.67
Percent Recovery 136.04 102.66 111.34
RSD 7.23 12.00 6.95
Diphenhydramine HCl Finished Product
Analysis 1 55.01 67.64 43.60
Analysis 2 62.65 51.09 51.80
Analysis 3 51.17 63.40 45.67
Average 56.28 60.71 47.02
Percent Recovery 112.55 121.42 94.04
RSD 10.38 14.16 9.07

Table 4. Discover SP-D Spike Recovery Results (ppb) of 50.0 ppb As, Cd, and Hg

Acetylsalicylic Acid Finished Product

Loratadine Finished Product

Diphenhydramine HCl Finished Product

  As Hg Cd
Acetylsalicylic Acid Finished Product
Analysis 1 56.16 52.53 53.50
Analysis 2 53.01 51.92 51.36
Analysis 3 55.53 46.90 50.65
Average 54.90 50.45 51.84
Percent Recovery 109.80 100.90 103.67
RSD 3.04 6.13 2.86
Loratadine Finished Product
Analysis 1 58.08 55.76 51.07
Analysis 2 54.09 54.79 51.34
Analysis 3 49.65 53.24 50.85
Average 53.94 54.60 51.09
Percent Recovery 107.87 109.19 102.18
RSD 7.82 2.33 0.48
Diphenhydramine HCl Finished Product
Analysis 1 66.23 53.78 61.64
Analysis 2 55.92 54.14 53.58
Analysis 3 55.99 53.19 56.12
Average 59.38 53.70 57.11
Percent Recovery 118.77 107.41 114.23
RSD 9.99 0.89 7.21

As shown in Table 3, MARS 6™ achieves good recovery of both volatile and non-volatile elements with cadmium at 94 – 112%, arsenic at 110 – 137%, and mercury at 101 – 122%. Moreover, the Discover SP-D also achieved good recoveries with 102 – 115% cadmium, 107 – 119% arsenic, and 100 – 110% mercury. These results show that the sample preparation with MARS 6™ and Discover SP-D meet the requirements of USP Chapter <233>.


Microwave closed vessel digestion instrumentation ensures rapid, convenient, and safe preparation of pharmaceutical samples, and hence it is ideal for metals analysis. MARS 6™ as well as Discover SP-D prepare pharmaceutical raw materials and finished products with respect to the sample throughput and laboratory workflow.

These instruments are suitable for a large scale operation and are less time-consuming. The chances of cross-contamination are eliminated as the systems employ completely sealed vessels. Further, the results of recoveries comply with the requirements of USP Chapter <233>.

About CEM Corporation

CEM is a company based on innovation that touches many different industries and scientific disciplines. It helped pioneer the field of microwave chemistry and has long been recognized for its expertise on the subject through publications and awards.

For more than 35 years, CEM has been designing and developing laboratory instruments and scientific methods (both microwave-based and non- microwave technologies) that are used by major companies, prestigious research institutes, and universities around the world.

CEM is the largest provider of microwave laboratory systems worldwide and has sold over 35,000 systems. The company also has the largest portfolio of microwave technology patents worldwide with over 300 patents. It has subsidiaries in the UK, France, Germany, Italy, and Japan and distributors in over 50 countries.

This information has been sourced, reviewed and adapted from materials provided by CEM Corporation.

For more information on this source, please visit CEM Corporation.


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