Siva Chaitanya Addala

PA/2015/102

Contents

• Introduction

• Speciation studies

• Different instrumental analytical techniques for elemental impurities

• References

Introduction

• Toxic metals like As, Pb, Cd, Hg, Se, Cr, Al, Ni, Cu and U enter the human body via the food chain including medicines, ambient air and drinking water leading to health problems.

• In addition, metal ions also can affect the stability and shelf life of the formulation, catalyze the degradation of the API’s leading to the formation of unqualified degradates

• Recently ICH has proposed safety standard guidelines for metal impurities (Q3D) for the purpose of quality assurance of pharmaceutical products

What is Speciation analysis ?

• Speciation analysis was first described in 1993 by Forstner

• The process of separation and quantification of different chemical forms of an element is more specifically termed speciation analysis.

• Inorganic As is much more toxic than the common organic forms, such as arsenobetaine, (the sum of arsenite (As(III)) and arsenate (As(V)) is below the limit.

• Similarly, the Hg limit is based on inorganic Hg (Hg2+), although methyl mercury (MeHg) is the more toxic form. The presence of MeHg in pharmaceuticals is considered unlikely, but it should be separated and measured specifically if samples are derived from material (for example, fish tissue) that may contain the compound in significant amounts.

• Distribution, Mobility, Bioavailability and Toxicity of trace metals in environmental and biological systems depend not simply on their concentrations, but critically on their chemical forms

• Individual metal species posses different chemical activity and ability to transform

• Speciation techniques using ICP-MS, ICP-AES, could be considered as the most sensitive and selectivetechniques

Different instrumental analytical techniques for elemental impurities

• Atomic absorption spectrometry (AAS)

• X-ray florescence spectrometry (XRF)

• Instrumental neutron activation analysis (INAA)

• Inductively coupled plasma atomic emission spectrometry (ICP-AES)

• Inductively coupled plasma mass spectrometry (ICP-MS)

Atomic absorption spectrometry (AAS)

• This technique is based on the principle that the amount of light absorbed is a measure of the concentration of a particular analyteat a particular wavelength

• GF-AAS is a technique which involves injection of a small amount of solution to be analyzed into a small graphite tube and thus is suitable for the analysis of metals at ultra-trace levels

• Mercury by ‘cold vapor’ method and some ‘volatile’ elements like As and Sb can be measured as their hydrides . A major advantage of cold vapor-AAS is the inherent separation of mercury from the matrix .

Both F-AAS and GF-AAS allow reliable determination of metallic impuritiesin pharmaceutical quality control operations.

Instrumental neutron activation analysis (INAA)

• INAA is a relatively straightforward analytical technique for determining elemental abundance in a wide range of materials.

• This technique relies on the measurement of characteristic radiation from radionuclides formed directly or indirectly by neutron irradiation of the material of interest. The energy of the emitted gamma rays is used to identify the nuclide and the intensity of the radiation can be used to determine its abundance.

• The advantages include,I. The method is non-destructive, hence the same sample can be

used for other measurements;II. Sample size can be very small, often as little as a milligram; III. Detection limits for many elements are in the ng/g range;IV. No sample preparation is required; and V. Over 40 elements can be measured simultaneously.

• Because of these advantages, INAA used to be a very popularanalytical technique compared to other analytical methods until ICPMS came in to use.

• As NAA does not require sample dissolution, it has a great advantage over solution techniques such as AAS, ICPAES and ICP-MS.

• Despite the above advantages, INAA is certainly not a popular analytical technique as it is time-consuming, not independent, requires a reactor nearby and involves longer cooling times for certain elements.

X-ray florescence spectrometry (XRF)

• In recent times XRF analysis has become increasingly attractive when compared to other techniques, especially due to the ease of sample preparation.

• XRF spectrometry involves irradiation of the samplewith high energy excitation X-rays and measurement of element-specific fluorescence X-rays at a particular wavelength or energy from the sample . Samples can be in solid, powder or liquid form.

• Since it is a non-contact analysis, problems such as memory effects commonly experienced in solution analysis, are not encountered.

• As it is a non-destructive technique, it is possible to reuse the sample after measurements.

• Both forms, namely, wavelength dispersive-XRF (WD-XRF) and energy dispersive-XRF (EDXRF) techniques have been successfully applied for the determination of Zn, Fe and Ni in API’s .

• Though there are several studies on the application of XRF techniques in pharmaceutical industry, because of the higher detection limits, they are not very popular for quantitativedeterminations of metal impurities in pharmaceutical samples.

Inductively Coupled Plasma

What is a Plasma?

•Plasma source provides atomization

•Plasma: “a gas-like phase of matter that consists of charged particles”

•ICP-AES plasma source is from the carrier gas

Typically argon is used

• ICP is a very powerful ionisation source

• Elements with IP < 8 eV are ionised to > 90%

• LODs for most metals are <ppb level

• Sample introduction is very versatile: liquid, gaseous or evensolid samples can be ionised in the Plasma

• The sample introduction allows a (relatively) straightforwardcoupling of chromatographic separation systems (GC, HPLC)

• ICP-MS detection is fast and multi-element capable

• A large variety of element speciation tasks can be tackled by GC or HPLC coupled to ICP-MS

• ICP-AES technique involves measurement of light emitted by the elements in a sample when introduced into an ICP source.

• The measured emission intensities are then compared to the intensities of standards of known concentrations to obtain the respective elemental concentrations in an unknown sample .

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Inductively Coupled Plasma Atomic Emission Spectrometry (ICP-AES)

• Limits of detection were at least a factor of ten below the USP limit concentrations confirming that the ICP-AES technique is well suited for quantitative determination of elemental impurities in pharmaceutical samples.

•The technique can simultaneously measure up to 60 elements with high sensitivity and an extraordinarily wide linear dynamic range which is perhaps the most outstanding feature of the ICP-AES

• The general chapter USP <233> includes two analytical procedures involving ICP-AES and ICP-MS for determination of elemental impurities in pharmaceuticals and includes a comprehensive validation procedure to ensure acceptability of results .

• Compared with F-AAS, ICP-AES provides lower detection limits, has multielement capability and a wider linear dynamic range

Inductively coupled plasma mass spectrometry (ICP-MS)

ICP-MS combines a high-temperature ICP source with a massspectrometer. The ICP source converts atoms of the elements in thesample to positively charged ions. These ions separated on the basisof mass-to-charge ratio in a mass spectrometer, are directed to adetector .

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• Further advances such as the advent of HR- ICP-MS and ICP-TOF-MS, during the last three decades have brought this technique to a point where this technique can deliver detection limits of one part in 1015 for a majority of elements in the periodic table.

• Detection limits are also far below the target limits in accordance with the analytical performance criteria described in USP 233. In fact.

• ICP-MS is one of the two spectroscopic methods which are included in the General chapter USP <233> for determination of elemental impurities in pharmaceuticals.

COST

Detection Limits• ICP-MS produces the best detection limits (typically 1-10 ppt)• Followed by GFAAS, (usually in the sub-ppb range) then ICP-AES

(of the order of 1-10 ppb) and finallyFAAS (in the sub-ppm range).

References

• V. Balaram ; Recent advances in the determination of elemental impurities in pharmaceuticals – Status, challenges and moving frontiers ; Trends in Analytical Chemistry