Extinction E

A few typical examples for the assay of pharmaceutical substances by UV-spectrophotometric method are described below  

Materials Required Amoxycillin trihydrate 0.17 g 100-ml volumetric flask 2 buffer solution pH 9.0 (Solution I Boric acid and Potassium Chloride (0.2 M)-Dissolve 12.366 g of Boric acid and 14.911 g of KC1 in DW and dilute with water to 1000 ml Solution II NaOH (0.2 N) Dissolve 8.0 g of NaOH in C02-free DW to produce 1000 ml Now, transfer 50 ml of solution I into a 200-ml volumetric flask 

Procedure Weigh accurately about 0.17 g of amoxycillin trihydrate and dissolve in sufficient DW to produce 500 ml. Now, transfer 10 ml of this solution into a 100 ml volumetric flask, add 10 ml of buffer solution pH 9.0 followed by 1 ml of acetic anhydride-dioxan solution, allow to stand for 5 minutes, and add sufficient water to produce 100 ml. Pipette 2 ml of the resulting solution into each of the two stoppered tubes. To tube 1 add 10 ml of imidazole-mercury reagent, mix, stopper the tube and immerse it in a water-bath previously maintained at 60 °C for exactly 25 minutes, with occasional swirling. Remove the tube from the water-bath and cool rapidly to 20 °C (Solution-1). To tube 2 add 10 ml of DW and mix thoroughly (Solution-2). Immediately, measure the extinctions of Solutions 1 and 2 at the maximum at about 325 nm, as detailed above, employing as the blank a mixture of 2 ml of DW and 10 ml of imidazole-mercury reagent for Solution-1 and simply DW for Solution-2. 

Calculations The content of C16H19N305S may be calculated from the difference between the extinctions of Solution-1 and that of Solution-2 and from the difference obtained by repeating the operation using 0.17 g of amoxycillin trihydrate (RS), instead of the sample being examined and the declared content of C16H19N305S in the amoxycillin trihydrate (RS). 

Cognate Assays Ampicillin can also be assayed by employing the above method using 0.15 g of the sample. 

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Any appropriate spectrophotometer capable for measuring both in the ultra-violet (UV) and visible range of the spectrum must essentially consist of an optical system that should produce monochromatic light in the range 190-780 nm and a suitable device for measuring the extinction (E) precisely and accurately. 

Theory Folic acid (I) undergoes cleavage by reduction with Zn-Hg in acidic medium to yield p-aminobenzoylglutamic acid (II). The primary aromatic amino group present in the latter is subsequently diazotized in the usual manner and coupled in acidic solution with N-(l-naphthyl)-ethylenediamine hydrochloride in the absence of light (caution). The colour thus produced has a maximum absorption at 550 nm and the extinction (E) is consequently compared with a calibration curve obtained from / -aminobcnzoic acid (PABA) that has been duly diazotized and coupled exactly in the same fashion as the/ -aminobcnzoylglutamic acid. 

As the inlet composition changes, HB points emerge from the first extinction E (a so-called Tokens-Bogdanov point) at 50% and 90% H2 in air. 

Before applying either the Lambert-Beer or the Biot law uncritically, one should check the range of linearity. As a rule of thumb, extinctions E above 1.5, but definitely those above 2.0, are to be discarded because linearity no longer holds. 

Table 3.1 summarizes observed absorption maxima as a function of various substituents. In particular, blue triphenodioxazines have very high molar extinctions e, comparable to those of bisazo and phthalocyanine dyes. Until recently, anthraquinone dyes (e ca. 15 000) were predominant in most applications requiring brilliant blue dyes, but the much stronger triphenodioxazine dyes now represent a less expensive alternative in many applications. 

Copper complexes of the formazan dye series (see Sections 2.10 and 3.11) are another alternative to reactive anthraquinone dyes they produce red to greenish-blue shades. Like triphenodioxazine dyes, copper formazans exhibit high molar extinctions ( e = 25 000 -30 000). These materials are derived from l-(2-hydroxyphe-nyl)-3-phenyl-5-(2-carboxyphenyl)formazan (22), in which all three rings are capable of supporting groups that increase the compound s reactivity and solubility. 

The structure of the paper itself affects the behavior with respect to Beer s law which establishes a relation between extinction E and concentration C of transparent solutions, according to the expression E = kC. When extinction measured on paper is plotted against concentration, not a linear but a hyperbolic curve is obtained (CIO, Cll). The cause of the error is the sievelike structure of the paper, since only the threads are covered with stained proteins while the meshes remain completely permeable to light (Fig. 34). This light falling directly on the photosensitive layer of the cell gives, on the microscopic scale, the same error as found for uneven distribution of stained spots on a clear... 

A suspension of 4.00g (6.75 mmol) of 3, 5 -bis-0-(p-nitrobenzoyl)-2 -deoxy-5-(trifluoromethyl)uridine in 250 ml of methanol was treated with 10 ml of diisopropylamine and refluxed until it had dissolved (about 18 minutes), and the solution was concentrated. The dry residue was partitioned between 50 ml of chloroform and 50 ml of water. The chloroform layer was washed with 20 ml of water, and the combined aqueous layers were concentrated. A low ultraviolet extinction (e 7200 and 262 pm pH 1) and the presence of isopropyl signals in the NMR spectrum (two singlets aty8.73 and 8.85) indicated the dry residue contained diisopropylamine, probably as a salt with the relatively acidic heterocyclic N-H in 14. 

As the pressure increases further, a second HB point (HB2) appears at the extinction point E and shifts toward the other HB (HB 1) point. An example is shown for 4 atm in Fig. 26.1c. Ignition /1 is no longer oscillatory, because the stationary partially ignited branch becomes locally stable in the vicinity of I. Time-dependent simulations indicate that the two HB points are supercritical, i.e., self-sustained oscillations die and emerge at these points with zero amplitude. In this case, the first extinction E defines again the actual extinction of the system. 

Infrared spectroscopy has not very often been used to check the deuterium purity of alkynes, with the exception of the case of compounds with the C=CD group. For these compounds a quantitative method, analogous to the one used to determine the deuterium content of deuterium oxide by n.m.r., has been described . Known quantities of the non-deuterated hydrocarbon RC=CH are successively added to the corresponding RC=CD compound of unknown deuterium purity, and the transmittance of the v(=CH) or K=CD) band is followed i>s. the added amounts of RC CH. The method is long but it does not require precise knowledge of the coefficient of molar extinction, e, of the v(=CH) band. Its main advantage is the elimination of errors due to molecular associations. It is not applicable to volatile compounds. 

Finally it should not be omitted to mention the fact that the rigid sphere fullerene, Ceo, can also be dissolved in vesicle membranes . When dissolved in hexane, chloroform or 1,2-dichloroethane, a narrow, concentration-independent absorption band at 334 nm (e 52000) was produced. In vesicles (lecithin, DODAB, DHP), the fullerene adsorption becomes concentration dependent whereby band-broadening, bathochromic shifts (343-360 nm) and loss of extinction (e 10000 4000) were observed in more concentrated solutions. 50 clearly aggregates within the vesicle membranes, a step not observed in micellar solutions. 

If the extinction (E) according to the Lambert-Beer law (see equation 2.4), which is calculated by equation (2.18), is less than 0.01,... 

Extinction E is the most frequently used measurable variable for light absorption. It is the decimal logarithm of the ratio of the intensity of the irradiated light (Iq) to the intensity of the light beam leaving the sample solution (I) ... 

The Lambert-Beer law states that light absorption (extinction E) is... 

By measuring the extinction E of a substance in solution of known concentration c, the molar extinction coefficient 8 may be determined ... 

The calibration curve provides information as to whether the Lambert-Beer law applies to the required concentration range, i.e. whether extinction E and concentration c are directly proportional. If this is the case, the calibration curve is a straight line. Deviations from the Lambert-Beer law in certain concentration ranges are indicated by the curved course of the calibration curve. 

Read off the manganese content from the calibration curve (C) on the basis of the measured extinction value (or if appropriate the net extinction E[ - E2 = E). Using the formula... 

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