X-Ray Fluorescence in Biological Sciences. Группа авторов

elements are determined using the standard calibration curve method, which is mostly used in cases where standards are unavailable.

      The ICP‐AES method is also used to determine the content of metals in wines and alcoholic beverages, heavy metals such as arsenic (As) in food stuffs, and trace elements in complex biomolecules such as proteins. It is capable to determine traces of oil additives which further indicate the service life left for the motor oil. It is capable to detect Li (Z = 3) to U (Z = 92) except gases, halogens, low contents of P and S. However in XRF, P and S can be easily determined and quantified. The combined use of ICP‐AES and ICP‐MS is very powerful and gives highly accurate and precise results for a broad range of elements from the major (percentage, %) to trace levels (typically sub ppb) [3].

      The disadvantage of this technique is that the emission spectra are complex and subject to spectral interferences for some elements. Matrix effects also create many challenges to quantifying the elements of interest. Some of the lither elements such as C, H, N, O, and halogens cannot be determined using this technique. Some elements cannot be detected by ICPs but often are subject to be analyzed exclusively by XRF such as S, Br, and Cl.

      In case of the limits of quantification equal to or above 1 ppm (μg/g), or where non‐destructive analysis is required, XRF is the most popular and attractive technique to analyze the solid samples, powders, oils, and slurry samples. As opposed to ICP‐AES, ICP‐MS, and AAS, it does not require sample dissolution or digestion and allows essentially non‐destructive analysis. In that case, XRF ensures accurate and reliable results by avoiding the potential for inaccuracies caused by incomplete dissolution and large dilutions.

      1.3.4 Ion Chromatography (IC)

      Ion Chromatograhy (IC) is a versatile analytical technique that is generally applied to detect positive and negative ions. It utilizes an ion's intrinsic affinity for both an “eluent” (typically buffered water) and a “stationary phase” (porous solid support with charge‐bearing functional groups) [3, 7]. It works on any kind of charged molecule, including large proteins, small nucleotides and amino acids.

There are two types of IC, cation‐exchange and anion‐exchange. Cation‐exchange chromatography is used for positively charged molecules (pH < pI). The stationary phase is negatively charged and positively charged molecules are loaded to get attracted toward it. In anion‐exchange chromatography, the stationary phase is positively charged and negatively charged molecules (pH > pI) are loaded to get attracted toward it. It is used in water analysis, protein purification, and quality control. Some of the ideal uses of IC include the analysis of food products and beverages, aqueous solutions such as water, and water‐extractable surfaces. It is more useful for analysis for quality assurance and control, purification of charged molecules, quantitative analysis of ions such as cations (Li, Na, K, Mg, Ca, NH₄ ⁺), anions (bromide, fluoride, nitrate, phosphate, sulfate, etc.), secondary amines, chlorite, sulfate, iodide, bromated, etc. Some of the advantages and limitations are tabulated in Table 1.2.

      IC has industrial applications too. Its main advantages in this sector include good precision and accuracy, reliability, high separation efficiency, high selectivity, good speed, and low operating cost. Applications of IC particularly in the field of pharmaceutical industry are being developed. These applications are typically focused on the determination of detection limits in the field of pharmaceuticals. The detection limits corresponding to oxalates, sulfamates, sulfates, iodide, phosphate, and electrolytes like sodium and potassium can be determined too. The IC can also be used for the analysis of drugs having pharmaceutical importance for the development of products with quality control testing. It can be used in pharmaceutical drugs in tablet or capsule form for the determination of the actual dosage of the drugs that can be dissolved within some time. IC can also be utilized for the detection as well as quantification of the inactive or undesirable ingredients that are being used in the pharmaceutical formulations. Sugar and associated alcohol have been detected in such formulations with the help of IC as they are easily resolved in an ion column due to having polar groups. IC can also analyze the impurities present in the drug substances and products. Impurities in the drug can be easily estimated and help to provide an intuition for the minimum and maximum dosage of drugs needed by a person on daily basis.

Table 1.2 Advantages and limitations of ion chromatography (IC) [3, 7].

Advantages	Limitations
Small sample quantity required.Rapid determination of anions and cations (inorganic as well as organic)Sensitivity: μg/l levelAnalysis of ionic speciesStability of the separator columns	Buffer requirementDetermination of only ionic analytesIdentification of peaks based on a retention time match to a standard solutionSmall change in pH greatly alters binding profile of stationary phase and ion statesSamples applied to the IC under conditions of low ionic strength and controlled pHResistant to salt‐induced corrosion

      1.3.5 Laser‐Induced Breakdown Spectroscopy (LIBS)

LIBS is a recently developed and rapid spectrochemical analysis technique and used to measure the relative elemental concentrations and their distribution within the samples [3, 5,15–18]. The schematic diagram of LIBS is shown in Figure 1.3. LIBS uses a high‐power pulsed laser beam that atomizes as well as excites the sample material. The creation of plasma happens only when the concentrated laser beam passes a threshold of power for an optical breakdown that is dependent of the target material and environment. LIBS can analyze any matter, be it solid, liquid or gas. Since all elements emit light of characteristic frequencies when excited to high temperatures, LIBS can detect all elements. It is limited by the power of the laser, sensitivity of the instrument, wavelengths corresponding to the spectrograph, and the detector. LIBS can be employed for the determination of the relative amount of element constituents and even impurities present in the sample material if the composition of the material is already known. Practically, it has been observed that the detection limits depend upon the temperature corresponding to the plasma excitation, the window used for the collection of light, and the line strength for the observed transitions.

      LIBS is more advantageous for depth analysis due to its refocused capability on the same location of a sample surface and its ability to provide depth profiling at a resolution of hundreds of nanometers per pass. In addition, the laser beam can be focused from 20 to 200 μm and thus allows the laser beam to scan across the whole sample surface that provides spatially resolved elemental mapping. It can detect nearly all the naturally occurring element down to down to ppm level, depending on the sample matrix.

Figure 1.3 A schematic setup of laser‐induced breakdown spectroscopy (LIBS).

LIBS can even detect halogen‐based agents. The detection of heavy and toxic elements such as lead (Pb) and mercury (Hg) in soil and plants can be determined by employing a field‐portable LIBS system. It has been observed that the analysis of the spectral emission of aluminum and aluminum oxides arises from the bulk aluminum in distinct bath gases can also be possible. It is used for kinetic modeling of LIBS plumes. It is also used to detect and discriminate various materials belonging to the category of explosives, geological, plastics, landmines, chemical as well as biological warfare agents.

      LIBS and XRF alike are generally used for positive material identification (PMI). For most of the applications, LIBS provides the same information as XRF, just using a laser source instead of radiation. But, in