What are the methods for studying biology? Biology methods

Refractometry is one of the simplest physical methods of analysis at a cost minimum quantity analyte and is carried out for a very short time. This method is used to identify substances, establish their purity, and determine the concentration of solutions.

The refractometry method is based on measuring the refractive index of light n by the analyte. The refractive index is the ratio of the speed of light in air to the speed of light in the substance under study. The value of the refractive index depends on the nature of the substance, temperature, and the wavelength of light at which the determination is carried out. In solutions, the refractive index also depends on the concentration of the solute and the nature of the solvent.

The different speed of propagation of a light beam in media with different densities causes a change in its direction when passing from one medium to another, i.e. refraction. The ratio of the speed of propagation of light in air v 1 to the speed of propagation of light in matter v 2, equal to the ratio of the sines of the angle of incidence of a ray of light α and the angle of its refraction β, is called index (coefficient) of refraction n and is a constant value for a given wavelength:

When a ray of light passes from a medium with a lower value of n to a medium with a higher refractive index (Fig. 13a) β< α. Если угол падения α луча С (рис.13б) приближается к 90 0 , то β < 90 0 . При дальнейшем увеличении угла падения (луч D) падающий свет полностью отражается от границы раздела и не попадает в менее плотную среду, происходит полное внутреннее отражение. Справа (при наблюдении против светового потока) от предельного луча D" находится затемненное поле, слева – освещенное поле.

Fig. 13. Refraction of a ray of light when passing from one medium to another:

a – refraction of a ray of light when passing from a less dense medium 1 to a more dense medium 2; b – refraction of a light beam at angles of incidence approaching 90 0; limit beam D - D" (total internal reflection).

The refractive index is determined using a special device called a refractometer. In practice, refractometers are used various systems: laboratory - RL, universal - RLU, RL - 2, "Karat - MT", etc.

The refractometer device is based on the phenomenon of total internal reflection of a light beam at the boundary of two media (one is a glass prism, the other is the analyzed solution) or on the position of the limiting beam at the border of light and shadow (Fig. 14).

Rice. 14. Diagram of the refractometer RL - 2:

1 – light from the source; 2 – mirror; 3 – lighting prism; 4 – measuring prism; 5 – compensator; 6- lens; 7 – prism; 8 – plate with crosshairs and a refractive index scale; 9 – eyepiece.

Light from source 1 hits mirror 2 and, being reflected, passes into the upper lighting prism 3, then into the lower measuring prism 4, made of special glass with a high refractive index. Between the hypotenuse surfaces of prisms 3 and 4, 1–2 drops of the analyzed liquid are placed using a capillary. To avoid mechanical damage to the prism, the capillary should not touch its surface.

The surface of the prism 4 serves as the interface at which the light beam is refracted. Due to the scattering of rays, the border of light and shadow turns out to be iridescent and blurry; dispersion compensator 5 eliminates this phenomenon. Next, the light passes through the lens 6 and the prism 7. On the plate 8 there are sighting lines (two crossed straight lines) and a scale of refractive indices observed in the eyepiece 9. The refractive index is read on the scale with three decimal places, the fourth digit is assessed by eye.

In eyepiece 9, a field with intersecting lines is visible to establish the interface. By moving the eyepiece, the crosshair point is aligned with the field interface (Fig. 15).

Rice. 15. Field of view in the refractometer eyepiece:

on the left – refractive index scale; on the right is a scale of percentage of dry substances; Between the prisms there is distilled water.

The position of the field interface corresponds to the angle of total internal reflection and depends on the refractive index of the analyzed liquid.

Laboratory refractometer RL - 2 (Fig. 16) has two scales - refractive index (from 1.33 to 1.54) and dry matter content, expressed in % (wt.) sucrose - from 0 to 95% (wt.) .

The refractive index is usually measured at a temperature of (20 ± 0.3) º C and a wavelength of the D line of the sodium spectrum (589.3 nm). The refractive index determined under such conditions is designated by the index n D 20.

The refractive index of distilled water is n 1 0 = 1.33299, practically the same index is taken as a reference as n 0 = 1.333.

Fig. 16. Refractometer RL – 2:

1 – base; 2 – column; 3 – body; 4 – dispersive dial to eliminate spectral coloring of light and shade; 5 – reflective mirror; 6 – chamber of the measuring prism; 7 – hinge connecting the prisms; 8 – lighting prism; 9 – thermometer; 10 – hole for adjusting the refractometer scale; 11 – scale for reading; 12 – handle; 13 – eyepiece

Operating procedure:

1. Checking the cleanliness of the contacting surfaces of the prisms (before starting measurements).

2. Checking the zero point. Apply 2-3 drops of distilled water to the surface of the measuring prism and carefully cover with the lighting prism. Open the lighting window and position it in the direction of the highest intensity of the light source using a mirror. By rotating the screws, obtain a sharp, clear distinction between light and dark fields in the field of view of the eyepiece. By rotating the screw, align the line of light and shadow exactly until it coincides with the point of intersection of the line in the upper window of the eyepiece. Vertical line in the lower window of the eyepiece indicates the measurement result - the refractive index of water at 20 ° C - 1.333. In the case of other readings, set the refractive index with a screw to 1.333, and use a key (remove the adjusting screw) to bring the boundary of light and shadow to the point of intersection of the lines.

3. Determination of refractive index. After installing the device to the zero point, lift the chamber of the lighting prism and remove the water with filter paper or a gauze cloth. Then 1-2 drops of the test solution are applied to the plane of the measuring prism, and the chamber is closed. Rotate the screws until the border of light and shadow coincides with the point of intersection of the lines. The refractive index of the solution is measured using a scale in the lower window of the eyepiece.

4. The relationship between the concentration of a two-component solution and the refractive index is established using a calibration graph. To construct a graph, standard solutions are prepared from a preparation of a chemically pure substance, the refractive indices are measured 3–4 times, the arithmetic mean is calculated and the resulting value is plotted on the ordinate axis, and the concentration of the solutions is plotted on the abscissa axis. Such a graph often represents an almost straight line. Having measured the refractive index of the analyzed solution, its concentration is found from the graph.

5. Finish work on the refractometer. After each determination, it is necessary to rinse both chambers with water and wipe dry with filter paper or a napkin; lay a thin layer of cotton wool between the chambers.

The refractive properties of a substance, determined by the structure of its molecule, are characterized by molecular refraction R m and are described by the Lorentz–Lorentz equation:

where M – molar mass substances, g/mol;

d – density x 10 3 kg/m 3.

Molecular refraction does not depend on temperature and state of aggregation substances. For chemical compounds it is an additive value that is used to establish the composition and structure organic matter. Molecular refraction is calculated as the sum of atomic refractions and increments of multiple bonds (Table 1). On the other hand, the refractive index and density of the identified substance are measured at 20 º C. These values, as well as the molar mass of the substance, are entered into the equation. In both cases, almost the same molecular refraction should be obtained.

Table 1

Atomic refractions of some chemical elements and increments of multiple bonds (20 0 C, λ = 589 nm)

Let's consider the calculation of molecular refraction using the example of chlorobenzene, the molecule of which contains 6 carbon atoms, 5 hydrogen atoms, 1 chlorine atom, and also has 3 double bonds, therefore:

R m= 6×2.418 + 5×1.100 + 1×5.967 + 3×1.733 = 31.2.

It is experimentally found that the refractive index of the analyzed liquid is 1.5248. The density of chlorobenzene is 1.107 × 10 3 kg/m 3, molar mass is 112.56 g/mol. We enter these values into the formula and get:

Slight difference between two values R m(31.2 – 30.9 = 0.3) indicate that the liquid being analyzed is indeed chlorobenzene. Significant discrepancies between the values of Rm found by the two methods may be due to experimental errors, significant contamination of the analyte, as well as the fact that the drug is not chlorobenzene.

Precautions during operation

Prisms in the device fail most quickly, so the following precautions must be observed when handling them.

1. Before determining the refractive index, the prisms are thoroughly cleaned of dirt and dust.

2. The refractive indices of acids and alkalis are not measured, since they corrode the surface of the prisms.

3. After taking measurements, wipe the surface of the prisms with a clean, soft cloth moistened with water or alcohol, wipe dry and place a small, dry, clean cloth or cotton wool between the prisms.

b) leave the test liquid between the prisms for a long time, since the surface of the prisms is then covered with a thin matte layer and measuring the refractive index becomes impossible.

Laboratory assignment No. 7

1. Determine the refractive indices of organic solvents and compare with known values n 20 D . Analyze the results obtained.

Organic solvents n 20 D

Ethanol 1.3613

Chloroform 1.4467

Toluene 1.4992

Methyl iodide 1.5207

Aniline 1.5863

1 – Bromonaphthalene 1.6582

2. Construct a calibration graph of the dependence of refractive indices on the concentration of ethyl alcohol in water.

3. Determine the concentration of the solution of ethyl alcohol in water given by the teacher.

4.Experimentally determine and calculate the molecular refraction of ethanol. Analyze the results obtained.

Laboratory work №8

The laws of physics play a very important role when carrying out calculations to plan a specific strategy for the production of any product or when drawing up a project for the construction of structures for various purposes. Many quantities are calculated, so measurements and calculations are made before planning work begins. For example, the refractive index of glass is equal to the ratio of the sine of the angle of incidence to the sine of the angle of refraction.

So first the process is underway measure the angles, then calculate their sine, and only then can you get the desired value. Despite the availability of tabular data, it is worth carrying out additional calculations each time, since reference books often use ideal conditions that can be achieved in real life almost impossible. Therefore, in reality, the indicator will necessarily differ from the table, and in some situations this is of fundamental importance.

Absolute indicator

The absolute refractive index depends on the brand of glass, since in practice there are a huge number of options that differ in composition and degree of transparency. On average it is 1.5 and fluctuates around this value by 0.2 in one direction or another. In rare cases, there may be deviations from this figure.

Again, if important exact indicator, then additional measurements are indispensable. But they also do not give a 100% reliable result, since the final value will be influenced by the position of the sun in the sky and cloudiness on the day of measurement. Fortunately, in 99.99% of cases it is enough to simply know that the refractive index of a material such as glass is greater than one and less than two, and all other tenths and hundredths do not matter.

On forums that help solve physics problems, the question often comes up: what is the refractive index of glass and diamond? Many people think that since these two substances are similar in appearance, then their properties should be approximately the same. But this is a misconception.

The maximum refraction of glass will be around 1.7, while for diamond this indicator reaches 2.42. Given gem is one of the few materials on Earth whose refractive index exceeds 2. This is due to its crystalline structure and the high level of scatter of light rays. The cut plays a minimal role in changes in the table value.

Relative indicator

The relative indicator for some environments can be characterized as follows:

- the refractive index of glass relative to water is approximately 1.18;
- the refractive index of the same material relative to air is equal to 1.5;
- refractive index relative to alcohol - 1.1.

Measurements of the indicator and calculations of the relative value are carried out according to a well-known algorithm. To find a relative parameter, you need to divide one table value by another. Or make experimental calculations for two environments, and then divide the data obtained. Such operations are often carried out in laboratory physics classes.

Determination of refractive index

Determining the refractive index of glass in practice is quite difficult, because high-precision instruments are required to measure the initial data. Any error will increase, since the calculation uses complex formulas that require the absence of errors.

In general, this coefficient shows how many times the speed of propagation of light rays slows down when passing through a certain obstacle. Therefore, it is typical only for transparent materials. The refractive index of gases is taken as the reference value, that is, as a unit. This was done so that it was possible to start from some value when making calculations.

If sunbeam falls on the surface of glass with a refractive index that is equal to the table value, then it can be changed in several ways:

1. Glue a film on top, whose refractive index will be higher than that of glass. This principle is used in car window tinting to improve passenger comfort and allow the driver to have a clearer view of traffic conditions. The film will also inhibit ultraviolet radiation.
2. Paint the glass with paint. This is what manufacturers of cheap products do sunglasses, but it is worth considering that this may be harmful to vision. IN good models The glass is immediately produced colored using a special technology.
3. Immerse the glass in some liquid. This is only useful for experiments.

If a ray of light passes from glass, then the refractive index is next material is calculated using a relative coefficient, which can be obtained by comparing table values. These calculations are very important in the design of optical systems that carry practical or experimental loads. Errors here are unacceptable, because they will lead to incorrect operation of the entire device, and then any data obtained with its help will be useless.

To determine the speed of light in glass with a refractive index, you need to divide the absolute value of the speed in a vacuum by the refractive index. Vacuum is used as a reference medium because refraction does not operate there due to the absence of any substances that could interfere with the smooth movement of light rays along a given path.

In any calculated indicators, the speed will be less than in the reference medium, since the refractive index is always greater than unity.

Lesson 25/III-1 Propagation of light in various media. Refraction of light at the interface between two media.

Learning new material.

Until now, we have considered the propagation of light in one medium, as usual - in air. Light can propagate in various media: move from one medium to another; At the points of incidence, the rays are not only reflected from the surface, but also partially pass through it. Such transitions cause many beautiful and interesting phenomena.

Changing the direction of propagation of light passing through the boundary of two media is called refraction of light.

Part of the light beam incident on the interface between two transparent media is reflected, and part passes into the other medium. In this case, the direction of the light beam that has passed into another medium changes. Therefore, the phenomenon is called refraction, and the ray is called refracted.

1 – incident beam

2 – reflected beam

3 – refracted ray α β

OO 1 – interface between two media

MN - perpendicular O O 1

The angle formed by the ray and a perpendicular to the interface between two media, lowered to the point of incidence of the ray, is called the angle of refraction γ (gamma).

Light in a vacuum travels at a speed of 300,000 km/s. In any medium, the speed of light is always less than in vacuum. Therefore, when light passes from one medium to another, its speed decreases and this causes the refraction of light. The lower the speed of light propagation in a given medium, the greater the optical density of this medium. For example, air has a higher optical density than vacuum, because the speed of light in air is slightly lower than in vacuum. The optical density of water is greater than the optical density of air because the speed of light in air is greater than in water.

The more the optical densities of two media differ, the more light is refracted at their interface. The more the speed of light changes at the interface between two media, the more it refracts.

For every transparent substance there is such an important physical characteristic, as the refractive index of light n. It shows how many times the speed of light in a given substance is less than in vacuum.

Refractive index of light

Substance	Substance	Substance
	Rock salt	Turpentine
	Cedar oil	Ethyl alcohol

Glycerol	Plexiglass	Glass (lightweight)
	Carbon disulfide

The ratio between the angle of incidence and the angle of refraction depends on the optical density of each medium. If a ray of light passes from a medium with a lower optical density to a medium with a higher optical density, then the angle of refraction will be less than the angle of incidence. If a ray of light comes from a medium with a higher optical density, then the angle of refraction will be smaller than the angle of incidence. If a ray of light passes from a medium with a higher optical density to a medium with a lower optical density, then the angle of refraction is greater than the angle of incidence.

That is, if n 1 γ; if n 1 >n 2 then α<γ.

Law of light refraction :

The incident beam, the refracted beam and the perpendicular to the interface between the two media at the point of incidence of the beam lie in the same plane.

The relationship between the angle of incidence and the angle of refraction is determined by the formula.

where is the sine of the angle of incidence and is the sine of the refraction angle.

The value of sines and tangents for angles 0 – 900

Degrees	Degrees	Degrees

The law of light refraction was first formulated by the Dutch astronomer and mathematician W. Snelius around 1626, a professor at Leiden University (1613).

For the 16th century, optics was an ultra-modern science. From a glass ball filled with water, which was used as a lens, a magnifying glass arose. And from it they invented a telescope and a microscope. At that time, the Netherlands needed telescopes to view the shore and escape from enemies in a timely manner. It was optics that ensured the success and reliability of navigation. Therefore, in the Netherlands, many scientists were interested in optics. Dutchman Skel Van Rooyen (Snelius) observed how a thin beam of light was reflected in the mirror. He measured the angle of incidence and the angle of reflection and established: the angle of reflection is equal to the angle of incidence. He also owns the laws of light reflection. He deduced the law of refraction of light.

Let's consider the law of light refraction.

It contains the relative refractive index of the second medium relative to the first, in the case when the second has a higher optical density. If light is refracted and passes through a medium with lower optical density, then α< γ, тогда

If the first medium is vacuum, then n 1 =1 then .

This indicator is called the absolute refractive index of the second medium:

where is the speed of light in a vacuum, the speed of light in a given medium.

A consequence of the refraction of light in the Earth's atmosphere is the fact that we see the Sun and stars slightly higher than their actual position. The refraction of light can explain the appearance of mirages, rainbows... the phenomenon of light refraction is the basis of the operating principle of numerical optical devices: microscope, telescope, camera.

Optics is a branch of physics that studies the nature of light radiation, its propagation and interaction with matter. Light waves are electromagnetic waves. The wavelength of light waves is contained in the interval. Waves of this range are perceived by the human eye.

Light travels along lines called rays. In the ray (or geometric) optics approximation, the finite wavelengths of light are neglected, assuming that λ→0. In many cases, geometric optics allows one to calculate the optical system quite well. The simplest optical system is a lens.

When studying the interference of light, it should be remembered that interference is observed only from coherent sources and that interference is associated with the redistribution of energy in space. Here it is important to be able to correctly write down the conditions for maximum and minimum light intensity and pay attention to issues such as the colors of thin films, stripes of equal thickness and equal inclination.

When studying the phenomenon of light diffraction, it is necessary to understand the Huygens-Fresnel principle, the Fresnel zone method, and understand how to describe the diffraction pattern on a single slit and on a diffraction grating.

When studying the phenomenon of polarization of light, you need to understand that the basis of this phenomenon is the transverseness of light waves. Attention should be paid to the methods of producing polarized light and to the laws of Brewster and Malus.