Quantification of group "B" vitamins by molecular spectroscopy in UV and visible regions in combined presence

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Quantification of group "B" vitamins by molecular spectroscopy in UV and visible regions in combined presence

Nickolay Vladimirovich Barykin,

Elena Viktorovna Cherdanseva,

Anatolii Ivanovich Matern

Abstract

Quantification of group "B" vitamins by molecular spectroscopy in UV and visible regions in combined presence

Nickolay Vladimirovich Barykin,¹ Elena Viktorovna Cherdanseva,²⁺ Anatolii Ivanovich Matern²*

¹ Department of Technology of Organic Synthesis;

² Department of Analytical Chemistry. Institute of Chemistry and Technology,

"Ural Federal University named after the first President of Russia B.N. Yeltsin", Mira Str., 19, Yekaterinburg, 620002. Russia, Тel.: 8(343) 375-97-56. Fax: 8(343) 375-61-44, E-mail: e.v.cherdantseva@ustu.ru.

In this paper, show how absorption spectra of substances vitamins solutions have been analyzed.

This data were ground for calculation of molar absorption coefficients.

We were detected a content (mole/dm³) of each component in a triple polyvitamin.

All in all, we were obtained following results:

· absolute error of mass content detection is 0.04-0.10 milligrams;

· relative error of concentration detection is 4.02-4.34 percent's.

Keyword: group "B" vitamins, quantification, molecular spectroscopy in UV and visible regions.

Introduction

Nowadays we live in the atmosphere of the constant stress and impairment ecology. It leads to the increase of the necessity of vitamins and polyvitamins. In one's turn, the production of provitamin's drugs needs in constant quality control. For this purpose highly sensitive method of the analysis is required. It enables to carry out the quantification of the vitamin in mixture. Vitamins B₂, B₆, B₉ are significant natural biologically active substance, which ensure normal work of human organism [1].

VITAMIN В₂

Riboflavin is a water-soluble vitamin has a flavin nature.

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For the first time, its crystal habit was extracted from whey and egg's proteins in 1933 by Richard Kuhn. In 1935 it was first synthesized.

Main functions of vitamin В ₂:

Ш participating in the building a number of enzymes, which takes part in the processes of energy exchange, synthesis of proteins and fat;

Ш forming a part of visual purple (rodopsin), which protects the retina from harmful influence of UV radiation.

Animal organism cannot synthesize riboflavin, that is why it is receive from meal or is synthesized by gut organisms.

Important sources of vitamin В ₂ are [2]: beef liver, kidney, heart, whole milk and milk dairy, yolk, yeast, grain crops and green vegetables.

VITAMIN В₆

Water-soluble vitamin is discovered by Paul Gyorgy (Paul Gyцrgy) in 1934.

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Pyridoxine Pyridoxal Pyridoxamine

Its crystal habit was isolated in 1938.

There are three individual substances, which possess the properties of vitamin В ₆:

All these forms can transform to each other, and the phosphorylated form of pyridoxal is the most biologically important.

Main functions of vitamin В ₆:

Ш forming a part of various enzymes and thus participating in metabolism of amino acids;

Ш having an effect on the metabolism of carbohydrates and fatty acids;

Ш more than 50 pyridoxalphosphate enzymes catalyzing various chemical reactions (amino acids decarboxylation; racemization; dehydrogenation; hydrolytic substrate decomposition).

Pyridoxine is received in organism from meal.

Important sources of vitamin В ₂ are [2]: meat, beef liver, kidney, brains, cod's caviar, yolk, milk, cereals and green vegetables.

VITAMIN В₉

(synonyms: folic acid, folacin, vitamin В_с)

It is a pterin ring fixed with glutamic acid residual through p-amino-benzoic acid residual.

Folacin is labeled as "water-soluble", but it dissolved in water badly. Folacin is dissolved in alkalescent solutions at room temperature and acid solutions when heated.

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In 1931 Lucy Wills published an article [3] covers the treatment of anemia by yeasts extract.

Main functions of vitamin В ₉ are determine by its participation in:

Ш enzymatic reactions;

Ш amino acids metabolism;

Ш purines and pyrimidines metabolism;

Ш process of hematogenesis and embryogenesis.

Important sources of vitamin В ₉ are [2]: fruits and green vegetables, strawberries, bread, beef liver.

Results and Discussion

There are characteristic absorption maxima for vitamins solutions at pH = 1 and these wavelengths, nm (Fig. 1): B_2-222, 263, 375 and 443; B_6-206 and 290; B_9-205 and 296.

Three wavelengths were selected after analyzing the spectra: with equal force absorb light (Fig. 1): B₂, and B₆ at 287 nm, B₂ and B₉ at 341 nm; at 375 nm exhibits a maximum absorption of B₂.

Fig. 1. The absorption spectrum of solutions: (a) - Vitamin B₂; (b) - vitamin B₆; (c) - Vitamin B₉

The experimental data are shown in Table. 1.

Tab. 1. Molar absorption coefficients of vitamins (е_i,j), and optical densities of triple polyvitamin solution (A) at the respective wavelengths

The absorption spectrum of a triple multivitamin solution at pH = 1 are shown in Fig. 2.

Fig. 2. The absorption spectrum of the solution triple multivitamin

Substituting the experimental data in the system (3) to give the system of equations (1):

A287= е(1,287)lc1+ е(2,287)lc2+ е(3,287)lc3

A341= е(1,341)lc1+ е(2,341)lc2+ е(3,341)lc3

A375= е(1,375)lc1+ е(2,375)lc2+ е(3,375)lc3 (1)

Indices 1, 2, 3 are designated as vitamins B₂, B₆, B₉.

For example, for the experiment number 1 (l = 1.000 cm):

1.93= 7500c₁ + 7600c₂ + 17200c₃

0.43= 5600c₁ + 5300c₃

0.33= 10300c₁ + 1300c₃

The results of the determination of the components in the triple polyvitamin are presented in Table. 2, 3.

Tab. 2. Mass determination of vitamins B₂, B₆, B₉ in polyvitamin

Tab. 3. Calculated concentration of component "i" in the polyvitamin solution (c_i); The relative error in determining the concentration of component "i" (д_i)

The relative error in the determination of concentrations of components multivitamin is 4 %, and the absolute error in the determination of the component masses is 0.04-0.10 mg.

Thus, the chosen method has shown good results (Table 2, 3) in determining the contents of the components in the ternary system.

Experimental

Molecular spectroscopy in UV and visible regions was selected as a method of quantitative analysis.

This method refers to middle-sensible and enables to determine a value of concentration in range 10^-6-10^-4 mole/md³ [4, 5]. Besides this method enables to determine a components concentrations in mixture, using the additivity of optical density on the basic law of absorption of electromagnetic radiation (Beer-Lambert-Bouguer law):

А_j = е_(i,j)•l•c_i (2)

A₁ = е₍₁,₁₎lc₁+ е₍₂,₁₎lc₂+...+ е_(n,₁₎lc_n

A₂ = е₍₁,₂₎lc₁+ е₍₂,₂₎lc₂+...+ е_(n,₂₎lc_n

A_m = е₍₁,_m)lc₁+ е₍₂,_m)lc₂+...+ е_(n,_m)lc_n, (3)

A_j - optical density of mixture at a wave-length "j";

е_(i,j) - absorption coefficirnt of component "i" at a wave-length "j", dm³/(cm•mole) or dm³/(cm•mg);

l - thickness of the absorption layer, cm;

c_i - concentration of component "i", mole/dm³ or mg/dm³.

The absorption spectra of the vitamins substances with a molar concentration of about 10^-5-10^-4 mole/dm³ were analyzed.

Hydrochloric acid (HCl) with a molar concentration of 1 mole/dm³ was used in the capacity of solvent.

The choice of solvent is due to the fact that in an acid medium:

Ш solubility of vitamin B₉ increases;

Ш studied vitamins are protonated forms [2], which are more stable.

PREPARATION OF SUBSTANCES VITAMINS SOLUTIONS

It is necessary get a substance's sample, calculated by the formula (4) to prepare a 100.00 cm³ of solution with a concentration of vitamin's substance numerically equal 10^-4 mole/dm³ in hydrochloric acid (the molar concentration of HCl is 0.1 mole/dm³).

m = c_i•V•M_i•10^-3, (4)

m - mass of substance's sample, g;

c_i - concentration of component "i", mole/dm³;

V - volume of solution, cm³;

M_i - molar mass of the component "i", g/mole.

Sample transfer into a volumetric flask Sample of vitamin B₉ must be taken in a glass-stopped flask, then add 10.00 cm³ of HCl solution (1 mol/dm³), close the stopper and dissolve when heated/ 100.00 cm³ quantitatively, add 10.00 cm³ of HCl solution (1 mole/dm³). Bring the volume in the flask to the mark by distilled water, mix thoroughly.

PREPARATION OF TRIPLE POLYVITAMIN SOLUTION

Take a sample of vitamin B₉ substance (2.0 mg) in a glass-stopped flask, add 10.00 cm³ of HCl solution (1 mole/dm³), close the stopper and dissolve when heated.

The resulting solution transfer into a volumetric flask 100.00 cm³ quantitatively.

Take a samples of substances of vitamins B₂ and B₆ (1.0 and 2.0 mg, respectively) transfer into the same flask quantitatively. Bring the volume in the flask to the mark by distilled water, mix thoroughly.

SPECTROPHOTOMETRIC STUDIES

Survey absorption spectra were carried out on a spectrophotometer SHIMADZU UV-mini 1240 quartz cells with a thickness of the absorbing layer l = 1.000 cm in the wavelength range from 190 to 540 nm.

A 33 survey of absorption spectrum of substances vitamins solutions (to 11 for each) were performed.

Based on the absorption spectra the molar absorption coefficients (еi, j) were calculated from the Beer-Lambert-Bouguer law (Table 1).

System of linear equations in three unknowns was derived from measurements of the optical density of the polyvitamin solution.

The concentrations (mole/dm³) of each component in the triple polyvitamin were determined by solving the system of equations by Cramer's method [6].

Math and statistical processing of data [4, 7] were performed using the program Microsoft Office Excel to optimize performance and save time.

Conclusion

Spectrophotometric analysis using the additivity of the optical density produces good results in the determination of low concentrations (10^-5-10^-4 mole/dm³) components of multivitamin medicines. In the first phase, the work was carried out with substances which are used directly to produce vitamins. This fact facilitates the adaptation of the analysis method to be used in production. The considered system is idealized: it contains all three components, while the real multivitamins contain six or more components. Possible improvement of the method, including by increasing the sample molar absorption coefficients of components and increasing the number of components in the system.

molar absorption vitamin spectrophotometric

References

[1] Komov V.P., Shvedova V.N. Biochemistry: a textbook for high schools. Moscow.: Drofa. 2004. 640 p.

[2] Zherebzov N.A., Popova T.N., Artuhov V.G. Biochemistry: a textbook. Voronezh: Voronezh State University Publishing. 2002. 696 p.

[3] Wills L. Treatment of "pernicious anaemia of pregnancy" and "tropical anaemia". BMJ. 1931. http://www.bmj.com/content/1/3676/1059.

[4] Zolotov U.A., Dorohova M.A., Fadeiva V.I. Fundamentals of Analytical Chemistry. T. 2. Methods of chemical analysis: a textbook for high schools. Moscow.: High school. 2004. 503 p.

[5] Bulatov M.I., Kalinkin I.P. A practical guide to the photometric and spectrophotometric methods of analysis. Leningrad.: Chemistry. 1976. 376 p.

[6] Pysmenyi D.T. Lectures on higher mathematics: a complete course. Moscow.: Iris-press. 2006. 608 p.

[7] Doerffel K. Statistics in Analytical Chemistry: translation from German. Moscow.: Mir. 1994. 268 p.

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