Chromatography Techniques in Drug Analysis

Introduction

Chromatography is a handy technique used in separation of concoctions that has been dissolved in a given liquid element. It is, therefore, referred to as a versatile skill of separating blended elements. Indeed, it uses a simple strategy of passing the soluble mixture over a substance that retains some specific elements dissolved in the original solution at different levels to form a feature commonly called a chromatogram. This procedure of separations operates under the perspective that different elements are retained differently given the same condition and exposure, which results to different elements flowing through the material at different rates.

Therefore, to undertake the whole process of separation using this method it is necessary to take a series of steps that encompass the use of various combination techniques and chemical components. Chromatography can be categorized into two major categories based on the procedural activities involved. These are sheet chromatography that involves the paper chromatography (PC) and the thin layer chromatography (TLC) as well as the Column chromatography that may involve Gas Chromatography (Poole 2003).

Analysis

In essence, chromatography is very important in two main ways: finding concentrations and chemical fingerprinting. In finding concentrations, chromatography includes a series of procedures. This may be well-illustrated in the gaseous chromatogram of gasoline and the ion chromatogram exhibited in orange juice. On the other hand, chromatography as a technique in chemical fingerprinting can be made in the identification of various species. In this regard, for instance, a harmful bee can easily be demarcated from the ordinary bee by contrast between the chromatogram of cuticle extorts. Furthermore, this procedure is also quite significant in tracing sources of contrabands and the successful detection of drugs in urine. This forms the basis upon which chromatography is used in the industry (Ettre & Hinshaw 2008).

Carbimazole and methimazole are types of drugs commonly used in the USA and the  UK respectively. The main treatment for which they are used is hyperthyroidism. Carbimazole is a yellowish-white powder with the chemical component Ethyl 3-methyl-2-thioxo-2-3dihydro-1H-imidazole. It is used in the thin layer chromatography as a control setup. The substance to be examined is dissolved in methylene chloride followed by successive dilution with the same solvent. Another setup of a solution of carbimazole is then dissolved in methylene chloride and diluted using the same solvent. The solution of carbimazole is set aside as a reference solution in the thin layer chromatography (TLC) under this setup. Then the development is closely observed under a path on the chromatography paper, 15 cm long and let to dry over a period of 30 minutes. This is then used to demonstrate a principal spot in the chromatogram under the scrutiny in ultra-violet light that appears on the same position as the one in the test solution, thus the comparative use of carbimazole in making a reference solution. Indeed, carbimazole contains some impurities identified in liquid chromatography involving dissolving it in acetonitrile and more than proportionate volume of water and comparing the structure of the chromatogram that in essence denotes impurities in carbimazole. These impurities include thiamazole and carbocisteine. Carbimazole has, however, some side effects despite their reputable usefulness in that it causes irritation in human body functioning, such as development of an infection, mouth ulcers disorderly bruising, and general body unfitness (Fried & Sherma 1999).

On the other hand, methimazole is a pale white crystalline powder with a boiling point of 280 ⁰c and a melting point of 146 ⁰c. This chemical substance is useful in making reference solution too during the test and treatment of hyperthyroid infections. In fact, carbimazole is a 3-carbethoxy extract of methimazole. However, despite the usefulness of methimazole, the chemical is also hazardous to some extent. For instance, health effects of methimazole may range from serious to less serious effects. Less serious effects include headache, drowsiness, and mild nausea, while serious effects include easy bleeding and unusual body weakness. Indeed, TLC is the processes used in separation of various components of benzodiazepines (BZD) chemicals, such as methimazole and its sub-constituent carbimazole, through simple procedure of dragenrdorff by thin layer chromatography (Hansen & Rasmussen 2012).

Paracetamol, also known as the ‘alvedon’, is another chemical component that can be determined through thin layer chromatography. It is a tablet found in a group of drugs known as analgesic. It is used to relieve mild to modest pains besides its usefulness in control of fever and related ailments. The chemical substance contains several components that are well analyzed in the paper chromatogram using the thin TLC method. As a matter of fact, when paracetamol is introduced in chromatography paper using thin layer chromatography, this can be used to determine its quality through identification of various components as laid out in the patterned chromatogram.

Paracetamol contains three major impurities as brought out in the chromatography process of European pharmacopoeia. These impurities are: 4-chloroacetanilide, 4-nitrophenol as well as 4-aminophenol all contained in the paracetamol and move in three distinctively different paces to form a distinct chromatogram when introduced in the chromatography paper. Three distinct impurities found in paracetamol are claimed to originate from synthesis. However, 4-aminophenol is the only impurity that is well-established to be genotoxic associated with degradation product and minimal in finished products. Indeed, the scrutiny of paracetamol under chromatographic impurities examination used in quality control and finger printing owing to its high sensitivity and specificity (Fritz & Gjerde 2009).

The main toxic impurity, 4-aminophenol, is an orthorhombic plate in water and a colorless crystal in physical appearance. However, upon exposure to light, the crystal may change to violet coloration. On the other hand, chloroacetanilide is a herbicide used in control of most annual grasses. Indeed, chloroacetanilide compounds undergo vigorous biodegradation processes when released into soil, thus mitigating their harmful effect that would result from failure to decompose.

Gas chromatography (GC) is used in column chromatogram where a sample mixture is introduced into a lengthy tubular column. Consequently, inert gas in a mobile phase sweeps through the tube pushing the mixture down the tube. As a result, stationary solid or syrupy liquid phase, which lines the tube, absorbs various components in the mixture selectively thus separating them into their constituent components. Often, the stationary phase is made of silicone polymers during gas chromatography (Cowper  & Derose 1983).

Finally, a specialized detector checks up on separated components leaving the tube. This method is often employed in analytical chemistry for separation and analysis of compounds that can be subjected to vaporization devoid of decomposition. These may include purity test and identification of compounds at times. GC is also a technique employed forensically in the process of drug analysis and arson toxicology. Indeed, this process of separation incorporates elementary properties of all methods of chromatography based on selective partitioning of particular compound amid different phases of specific substance under scrutiny. The mobile phase is an inert gas, such as helium, while the other phase comprises of a waxy substance (stationary phase) encrusted on a solid inside the chromatographic column (Pecsok & Drew 2009).

Chloroxylenol used in gas chromatography is a highly toxic substance to various bacteria and fungi but has mild side effects on human body performance. Consequently, it is used for application in antiseptics in human environment, including the dettol. Indeed, chloroxylenol assumes very low systemic toxicity and has very little if any effect even when used over the recommended dosage in dettol. On the other hand, chlorocresol is a chemical substance, with is  a colorless crystal at a normal room temperature. The substance is also vaguely soluble in water and it often dissolved in alcohol for use as a disinfectant as the case of dettol. Indeed, it is a chlorine saturated phenol, which is used in industries as an antiseptic often in line with dettol detergent identified by means of chromatography. Furthermore, the chemical substance is used as a preservative of its counterpart product, in this case, the dettol. However, antiseptic has some side effects in that it is moderately allergic to sensitive skins.

Essential composition of volatile oils was classified by GC-FID as well as GC-MS. During this process, chromatography was used to identify 73 and 70 composites respectively. This represented above 90.3 and 98.7 per cent of oil composition in the category G Sylvaticum. Additionally, sesquiterpene hydrocarbons also emerged as the main constituent in the analysis. Furthermore, the analysis showed that muurolene was the main component of oils in G. Sylvaticum. In summation, comparative experimentation unveiled the irrefutable fact that level of aggregate volatile (98.7 per cent) and the main components (39.8 per cent) were relatively good in MD G. Sylvaticum (Moldoveanu & David 2002).

The head space gas chromatography is a technique used in relaying volatile components of liquids and solid substances and mounting them onto gas chromatograph. ‘Head space’ refers to the space on the upper face of the liquid or solid sample, on which a sealed vial is placed. The instrument used removes a reducible value of headspace and mounts it to the carrier gas to gas chromatograph. This process is used in the analysis of both fragrances and consumer products.  A night nurse liquid is an absolute night time cure for flu and cold, thus relieving tickly cough and shivers. However, this drug contains paracetamol and promethazines, which are identifiable in the chromatogram and are suitable for adults (Quintin 2010).

Indeed, the night time syrup contains liquid ethanol. In essence, it contains alcohol, which is vital in medication as it contains preservative features and acts as a solvent to various elementary components of liquid syrup. According to naturopathic expert Francis Binker, ethanol is a good solvent for various ingredients that water may not dissolve, thus preventing settling of such at the bottom of the container. However, alcohol has some side effects in taking such medication. For instance, a frequent use of alcohol, especially in combination with acetaminophen, leads to an increased affinity for liver damage and may also result in addiction (Quintin 2010).

Paracetamol tablet has been determined in components and features by use of HPLC method. Indeed, this method has been used in separation of paracetamol, caffeine, and dipyrone using ‘Bond pack’ isocratic elution. Furthermore, method is linear and reproducible chromatographic procedure that sees the use of paper medium in separation of the components above due to their differences in adhesive capabilities. As pointed out earlier, paracetamol tablet contains three major impurities, namely 4-nitrophenol, 4-aminophenol as well as 4-chloroacetanilide. However, only 4-aminophenol impurity is quite toxic to human health (Cazes & Scott 2002).

Introduction of corticosteroid will see formation of an alignment of rings on the medium (paper) denoting the successive separation of various elements making up the sample mixture of corticosteroids. These corticosteroids are main nine in numbers and are effectively separated using the HPLC method. They include hydrocortisone, cortisone, prednisone, and prednisolone. Hydrocortisone is a gentle corticosteroid used in moisturizing inflammatory skins, for instance, the eczema flare ups. It relieves flare-up related symptoms thus relieving inflammations resulting from irritants such as insect stings and allergens. The drug has minimal if any side effects. However, for long time users, there may be development of permanent ‘striae’ features, which are stretch-like marks as well as bruising and discolored constricted blood vessels (Cazes 2010).

On the other hand, cortisone prednisone is also a form of steroid produced by the natural mechanism of adrenal glands when the body is stressed. This natural form of cortisone is released into blood but has minimal reactions. However, the injectable form of cortisone is used to relieve inflammation and does not mitigate pain. However, cortisone injection may result in a number of side effects such as ‘cortisone flares’ and joint infection. Flare infection occurs when a given injection of cortisone crystallizes in the body resulting in a short-term period of pain. Finally, prednisolone is the other type of steroid that serves to eliminate inflammation causing micro-organisms released by certain cells of the body, thus plummeting inflammation. Indeed, prednisolone is a synthetic glucocorticoid used in replacement therapy as an anti-allergic drug. It is a white and crystalline powderly substance (Moldoveanu & David 2002).

High performance liquid chromatography has been quite effective for profile psychoanalysis of steroids. HPLC has been quite effective since high temperature condition of the water used is a modest polarity solvent. Additionally, due to high temperatures, there is reduction in viscosity of the mobile phase, thus lower back pressure, higher rates of flowing, and short analysis time. This mounts to increased efficiency. Furthermore, HPLC has relatively high efficiency due to superior diffusion rates that improve sensitivity and resolution (Boyer & Tsao 1993).

Finally, HPLC has been the most outstanding method of separating and determining dichlofenac found in biological solutions. Direct injection and protein precipitation has been some of the procedures that have been used to describe analysis of dichlofenac sodium. Other processes used include solid-phase extractions (SPE) among others. Indeed, this compound is used as anti-inflammatory analgesic drug for the purpose of relieving pain as well as treating rheumatic infections. The drug is used in the treatment of rheumatoid arthritis. Dichlofenac is an off-white crystalline solid in nature also used as a cyclo-oxygenase inhibitor. Additionally, it is easily dissolved in methanol, ethanol, and slightly soluble in acetone and water. However, despite the positive implication of dichlofenac acetate, it has some side effects, such as impaired breathing, swelling tongue, throat or lips, as well as hives may occur with successive use of dichlofenac. Synthetic precursor is one of the impurities of dichlofenac drug. The Reverse-Phase HIGH Performance Liquid Chromatographic (RP-HPLC) technique is preferably quite effective in determining the impurities since it evaluates aceclofenac and other related impurities simultaneously. This leads to increased efficiency and convenience in carrying out the task of identification and the separation of impurities as well (Ettre & Hinshaw 2008).

In conclusion, chromatography is a very important procedure with broad usage particularly in pharmaceutical industry. This process assists in separation of various elements of drugs as well as impurities therein. Consequently, it is used in purifying various compounds of drugs and related components. In essence, chromatography operates under the broad principle of differences in power of dissolving as well as the pace at which different substance fuse through a given medium, often paper. Indeed, this analytical method is quite useful in inspection of industrial pharmaceutical products.