ICP-MS: Measuring Phosphorus and Sulfur in Organic Matrices

News & Events At Solvias, we see challenges as a chance to grow. Elements like phosphorous and sulfur require special treatment in order to be measured with accuracy using inductively coupled plasma mass spectrometry, ICP-MS. Part of the treatment is to remove interferences like ions (from the gas or acid used, and/or from other elements present in...

News & Events

At Solvias, we see challenges as a chance to grow. Elements like phosphorous and sulfur require special treatment in order to be measured with accuracy using inductively coupled plasma mass spectrometry, ICP-MS. Part of the treatment is to remove interferences like ions (from the gas or acid used, and/or from other elements present in the sample) which combine to form a mass corresponding to the analyte of interest.

Why measure sulfur and phosphorus?

Phosphorus and sulfur are extensively used in the production of drugs, especially biologics. With biological molecules becoming more prevalent in the pharmaceutical industry, so is the presence of phosphorus and sulfur in drug products.

Phosphorus is an integral part of the "backbone" of nucleotides and thus of all short fragments of DNA and RNA, so it can be used to quantify such molecules. Sulfur has long been present in small and large molecules, either as an active group, e.g. in thiols, or within the normal structure of the molecule, for example, in penicillin, and in some cases also as a disulfide. In biologics, sulfur occurs frequently in peptides and proteins in the amino acids cysteine and methionine. Both phosphorus and sulfur are irreplaceable in various applications and at the same time may act as unwanted contaminants. Consequently, it is essential to quantify the elemental concentrations in a residue.

ICP-MS is a state-of-the-art element quantification technique that has been commonly used in recent years in the pharmaceutical industry. The implementation of this analytical technique in the ICH Q3D "Elemental Impurities" has spread its fame that ICP-MS is THE ideal method to determine element concentrations in the low concentration range.

Measuring phosphorus and sulfur by ICP-MS – O2 reaction cell.

Phosphorus is a monoisotopic element of mass 30.97 amu (reference: https://pubchem.ncbi.nlm.nih.gov/compound/5462309#section=Chemical-and-Physical-Properties). For a mass-based analytical technique like ICP-MS, this means that only the one isotope (31P) can be selected to quantify phosphorus. Sulfur, on the other hand, has three naturally stable isotopes (32S, 33S, 34S), with an abundance of approximately 95% for 32S. For the determination of trace concentrations, the only reasonable isotope is presented by 32S.

In ICP-MS analyses the introduced sample is largely ionized in the plasma followed by a separation of the ions by their mass-to-charge ratio. A resulting ion beam is then quantified via calibration of the associated signal intensities. Since the argon plasma is operated under atmospheric conditions and the analytes are transported there either in acidified (or basic) aqueous solution or in organic solvents, the most abundant elements in the plasma next to Ar are H, O, N, and C. Thus, the low mass-range (correctly, the low mass-to-charge ratio range) of the ICP-MS is very "crowded" by NO+, O2+, CO+, varying in their mass by the combination and abundance of their respective parts. For example, 16O2+ on m/z 32 is very common, and while 15O2+ on m/z 30 is much less abundant, 14N16O+ also contributes there. In case the mass of interest is affected by such polyatomic interferences, we need a workaround to find an effective way to solve the problem. For oxide-forming analytes, this can be achieved elegantly in a reaction cell by the means of oxygen.

While passing through the reaction cell, sulfur and phosphorous ions are forced to interact with oxygen molecules, which in the event of oxidation is associated with a mass shift. When the interfering ions (e.g. 16O2 and NO+) are not equally oxidized, the interference can efficiently be removed. Examples of the mass shift for sulfur and phosphorous induced by oxygen are shown below:

For sulfur, which receives strong interference from the ion 16O2+:
mass shift from 32 to 48 amu:

32S+ + 16O2 → reaction cell → 32S16O+ + 16O

Behavior of the interfering ion:
mass remains 32 amu:

16O2+ + O2 → reaction cell → no reaction

For phosphorus, which receives strong interference from multiple ions (like 15N16O+, 1H14N16O+, 13C18O+, ...):
mass shift from 31 to 47 amu:

31P+ + 16O2 → reaction cell → 31P 16O+ + 16O

For the interfering species, mass remains 31 amu:

15N16O+ + O2 →reaction cell → no reaction

In both cases, mass shifting using O2 as a reaction gas resolves the issue of polyatomic interferences in the determination of trace concentrations of sulfur and phosphorous.

However, it must be kept in mind that although the analyte can then be analyzed at a mass different to its interfering species, the mass it is shifted to could be already be "occupied" by other analytes also present in the sample, e.g. 48Ti in the case of 32S16O or 47Ti in the case of 31P 16O. In such cases, with a normal single quadrupole ICP-MS system, one would end up with the same problem, just at another mass.

Benefit of triple quadrupole ICP-MS

A triple quadrupole allows, compared to simple quadrupole ICPMS, a finer analysis due to the addition of a supplementary quadrupole to the system before the collision / reaction cell. This addition acts as a supplementary mass filtering unit thus removing more of the interferences.

Since the quadrupoles can be tuned to select for different masses, they are ideally suited for the above-listed interference. The first mass filter is then set to the analyte plus its interference, e.g. m/z 32 for sulfur, then after the reaction cell (which features the second quadrupole), the third quadrupole then is set to m/z 48. Thus, any interference which may have been present in the sample at the mass to be shifted to, has already been removed by the first mass filter.

A little intensity is lost through the process, but the gain is worth the loss: interference being drastically lowered, the collision/reaction cell's performance is at its best level.

Advantages and disadvantages of this technique

ICP-MS is a non-species selective analysis, in the sense that only the total amount of a specific element is measured and not its oxidation state. For instance, this means that it isn't possible to know whether the sample contains sulfur in multiple molecular forms; only total sulfur content is determined. The species-selective quantification of sulfur, sulfate or sulfur in a free form for instance is only possible by coupling ICP-MS to a separation technique like HPLC-ICP-MS.

On the other hand, in a known matrix with a known analyte, a very accurate quantitation can be performed by the mean of a conversion factor for calculation of the species of interest, measuring e.g. total phosphorus to express the results in phosphate.

The advantage of this method is the principle of analysis itself that gets rid of organic compounds that could interfere in other types of measurements by a digestion step (solid samples) and/or directly by the subsequent plasma ionization. Nevertheless, one restriction applies to volatile sulfur species due to the sample preparation: in the process of mineralization, volatile sulfur species are lost.

At Solvias, we are currently validating a general method with an investigated limit of quantitation targeted at 0.5 mg/kg for both phosphorus and sulfur, aimed mainly at organic matrices, whether heavier or lighter (such as H2O2 used in electronical circuit cleaning)

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Source: www.solvias.com