1. A sample application pen for preparing thin-layer chromatography is characterized by comprising a liquid suction head, a liquid storage cavity, a sample application pen point and a filling pen core; the liquid suction head is communicated with the liquid storage cavity; the liquid storage cavity is detachably connected with the sample application pen point; the filling pen core is filled in the sample application pen head.

2. The printer for thin layer chromatography according to claim 1, wherein the pipette tip is hollow inside; the liquid suction head is an elastic liquid suction head, can generate elastic deformation in an extrusion state, and can recover the original shape after the extrusion force disappears.

3. The printer pen for preparing thin layer chromatography as claimed in claim 2, wherein the pipette tip is made of one or more of latex, silica gel, rubber, and plastic.

4. The printer for thin layer chromatography as claimed in claim 2, wherein the pipette tip is further provided with an opening having a plug with a size matching with the size of the opening.

5. The spotting pen for preparing thin-layer chromatography according to claim 1, wherein the outer periphery of the reservoir chamber is provided with a protective jacket; the material of the liquid storage cavity is more than one of plastic, glass, latex, silica gel and rubber; the material of the point-application pen head is more than one of plastics, glass, latex, silica gel and rubber.

6. The spotting pen for preparing thin-layer chromatography as claimed in claim 1, wherein the top of the spotting pen head is provided with a port matching with the bottom of the reservoir, and the bottom of the reservoir is hermetically connected with the spotting pen head through the top opening into which the spotting pen head is inserted.

7. The printer for thin layer chromatography as claimed in claim 1, wherein the printer tip is provided with an opening at the bottom, and the filling cartridge passes through the opening and extends to the outside.

8. The spotting pen for preparing thin-layer chromatography according to claim 1 characterized in that the top of the filling cartridge is in contact with the bottom of the reservoir when the reservoir is connected to the spotting pen tip.

9. The spotting pen for preparing thin-layer chromatography as claimed in claim 1, wherein the material of the filling pen core is more than one of fiber, cotton, hair, silk thread.

10. Use of a printer for thin layer chromatography according to any of claims 1 to 9 for thin layer chromatography printing, comprising the steps of:

(1) extruding the liquid suction head to make the liquid suction head generate elastic deformation, inserting the bottom of the liquid storage cavity into the liquid to be spotted, stopping extruding the liquid suction head to make the liquid suction head restore to the original shape, and thus making the liquid storage cavity absorb the liquid to be spotted;

(2) and (2) connecting the liquid storage cavity for absorbing the liquid to be spotted in the step (1) with the spotting pen point, placing the pen point of the spotting pen point on the position needing spotting, contacting the bottom of the filling pen core with the position needing spotting, soaking the liquid to be spotted into the filling pen core, and applying the liquid to the position needing spotting through the filling pen core to finish spotting.

Background

Natural products or organic synthetic products often involve the isolation and purification of a sample. The thin-layer chromatography is an important separation method and means in the field of natural products, and especially plays a key role in preparation of trace components with remarkable activity. The efficient preparative chromatography technology not only saves the time for separation and purification, but also can obtain a compound with a novel structure, and is beneficial to promoting the rapid development of the research field of natural drugs. It is important to note that the loading process is an important and critical step in the preparative thin layer chromatography technique. However, at present, the sample is applied on the thin-layer plate successively and repeatedly by dipping the sample liquid in the glass capillary tube, which has the restrictive problems of poor sample application effect, low efficiency and long time consumption.

Disclosure of Invention

In order to overcome the defects of the prior art, the invention aims to provide a spotting pen for preparing thin-layer chromatography and application thereof.

The invention aims to provide a thin-layer chromatography sample application pen. The sample application pen is simple and convenient to operate and easy to control, and can improve the working efficiency and sample application effect of sample application on a prepared thin-layer plate.

The purpose of the invention is realized by at least one of the following technical solutions.

The invention provides a sample application pen for preparing thin-layer chromatography, which comprises a liquid suction head, a liquid storage cavity, a sample application pen point and a filling pen core; the liquid suction head is communicated with the liquid storage cavity (the liquid suction head and the liquid storage cavity are in sealed connection to ensure air tightness); the liquid storage cavity is detachably connected with the sample application pen point; the filling pen core is filled in the sample application pen head.

Further, the interior of the liquid suction head is hollow; the liquid suction head is an elastic liquid suction head, can generate elastic deformation in an extrusion state, and can recover the original shape after the extrusion force disappears. The sample liquid can be absorbed by extruding the liquid suction head and stored in the liquid storage cavity.

When the liquid suction head is extruded, the air in the liquid suction head is discharged, when the extrusion force disappears, the liquid suction head restores to the original shape, negative pressure is formed in the liquid suction head, and under the action of atmospheric pressure, the liquid to be spotted is sucked into the liquid storage cavity.

Preferably, the material of the liquid suction head is one or more of latex, silica gel, plastic and rubber, but is not limited to the above materials.

Preferably, the liquid suction head is also provided with an opening, and the opening is provided with a plug (air valve plug) with matched size. The open pore can play a role in balancing air pressure, sample liquid is sucked in the state that the air valve plug is closed, and after the liquid is sucked, the plug can be opened, so that the air pressure inside and outside the liquid suction head is balanced, and the sample liquid permeates into the sample dispensing pen core. The invention can adjust the internal and external air pressure of the pipette head and the liquid storage cavity by closing or opening the plug of the opening on the pipette head, thereby keeping or releasing the sample solution in the liquid storage cavity.

When the viscosity of the liquid to be spotted is small or the density is large, the plug can not be pulled out, the speed of the liquid to be spotted permeating into the filling pen core is not accelerated, and the spotting is directly finished. The invention can adjust the balance of the internal and external air pressures of the liquid suction head by pulling out the plug or not, further can control the speed of the liquid to be spotted penetrating into the filling pen core, and finally controls the spotting speed, so that the sample loading effect is more uniform, thereby being suitable for experiments limited by various conditions.

When the liquid suction head is not provided with the opening, the liquid in the liquid storage cavity can be contacted with the filling pen core by extruding the liquid suction head, the liquid to be subjected to sample application is soaked into the filling pen core, and the liquid is applied to a position needing sample application through the filling pen core to finish sample application.

Further preferably, the opening may be provided at the top of the pipette head.

Furthermore, a protective layer is arranged around the liquid storage cavity; the material of the liquid storage cavity is one of plastic, glass, latex, silica gel and rubber, but is not limited to the above materials.

Further, the spotting pen head is made of one of plastic, glass, latex, silica gel and rubber, but not limited to the above materials.

Furthermore, the bottom of the sample application pen point is provided with an opening, the top of the sample application pen point is provided with a connector matched with the size of the bottom of the liquid storage cavity, and the sample application pen point can be hermetically connected with the connector at the bottom of the liquid storage cavity when the connector at the bottom of the liquid storage cavity is inserted into the connector at the top of the sample application pen point.

Preferably, the bottom of the filling cartridge passes through the opening of the printing pen tip and extends to the outside.

Further, when the liquid storage cavity is connected with the sample application pen point, the top of the filling pen core is connected with the bottom of the liquid storage cavity.

Further, the material of the refill is one or more of fiber, cotton, hair and silk thread, but is not limited to the above materials.

The application of the sample application pen for preparing the thin-layer chromatography provided by the invention in the sample application of the thin-layer chromatography comprises the following steps:

(1) extruding the liquid suction head to make the liquid suction head generate elastic deformation, inserting the bottom of the liquid storage cavity into the liquid to be spotted, stopping extruding the liquid suction head to make the liquid suction head restore to the original shape, and thus making the liquid storage cavity absorb the liquid to be spotted;

(2) and (2) connecting the liquid storage cavity for absorbing the liquid to be spotted in the step (1) with the spotting pen point, placing the pen point of the spotting pen point on the position needing spotting, contacting the bottom of the filling pen core with the position needing spotting, soaking the liquid to be spotted into the filling pen core, and applying the liquid to the position needing spotting through the filling pen core to finish spotting.

Preferably, when the liquid suction head is provided with an opening and a gas valve plug, the application of the spotting pen for preparing thin-layer chromatography provided by the invention in thin-layer chromatography spotting comprises the following steps:

(1) closing an air valve plug on the liquid suction head, extruding the liquid suction head to enable the liquid suction head to generate elastic deformation, inserting the bottom of the liquid storage cavity into the liquid to be spotted, and stopping extruding the liquid suction head to enable the liquid suction head to recover the original shape, so that the liquid to be spotted is absorbed by the liquid storage cavity;

(2) and (2) connecting the liquid storage cavity for absorbing the liquid to be spotted in the step (1) with the spotting pen point, opening the air valve plug at the top of the liquid suction head to balance the internal and external air pressures, soaking the liquid to be spotted in the pen core, and applying the liquid to the position required by preparing the thin-layer chromatography through the filling pen core to finish sampling.

Further, in the step (1), before the liquid suction head with the air valve is pressed, the plug needs to be closed, and then the pressing and the subsequent liquid suction work are carried out.

Further, in the step (2), after the bottom of the filling pen core is contacted with the position needing sample application, the air valve plug can be opened to balance the air pressure inside and outside the liquid suction head, the liquid to be subjected to sample application is soaked into the filling pen core, and the liquid is applied to the position needing sample application through the filling pen core to complete sample application.

The invention finds that negative pressure is easily formed in the closed liquid suction head, so that the liquid to be spotted cannot contact with the filling pen core and cannot be spotted. Therefore, in the preferable scheme of the invention, the liquid suction head is provided with the opening and the plug (air valve plug) with the corresponding size, the invention can adjust the internal and external air pressure of the liquid suction head by opening or closing the air valve plug at any time, which is not only beneficial to the liquid to be spotted permeating into the filling pen core, but also avoids the liquid to be spotted directly dripping from the filling pen core.

Compared with the prior art, the invention has the following advantages and beneficial effects:

(1) the sample application pen for preparing the thin-layer chromatography provided by the invention has the advantages of simple and reasonable structure and simple and convenient operation. The working efficiency and the sample effect of sample application on the preparation of the thin-layer plate can be improved; the sample application pen has the characteristics of one-time sample application, short sample application time and the like, and has the remarkable advantages of small damage to a preparation plate, accurate sample application, unified standard and the like in the sample application process.

(2) The prepared thin-layer chromatography sample application pen provided by the invention solves the problems of thin-layer surface damage, uneven sample application and the like caused by multiple times of sample application, long time, multiple times of sample liquid absorption and multiple repeated sample application by using a conventional glass capillary; the product of the invention has simple manufacturing process, can simplify the experimental operation steps and can be repeatedly used.

Drawings

FIG. 1 is a schematic diagram of a configuration of a spotting pen for preparing thin-layer chromatography provided in example 1 of the present invention;

wherein, the liquid suction head 1, the liquid storage cavity 2, the point sample pen head 3 and the filling pen core 4 are arranged;

FIG. 2 is a graph showing the results of thin layer surface breakage after loading of a spotting pen and a capillary tube for thin layer chromatography in example 3 of the present invention;

FIG. 3 is a graph showing the results of spotting the surface of thin layers on a spotting pen and capillary tubes for thin layer chromatography prepared as described in example 4 of the present invention after loading.

Detailed Description

The following examples further illustrate the practice of the present invention, but are not intended to limit the practice or protection of the invention. It is noted that the processes described below, if not specifically described in detail, are all realizable or understandable by those skilled in the art with reference to the prior art. The reagents or apparatus used are not indicated to the manufacturer, and are considered to be conventional products available by commercial purchase.

Example 1

Referring to fig. 1, a spotting pen for preparing thin layer chromatography comprises a liquid suction head 1, a liquid storage cavity 2, a spotting pen head 3 and a filling pen core 4; the liquid suction head 1 is communicated with the liquid storage cavity 2 (hermetically connected to ensure air tightness); the liquid storage cavity 2 is detachably connected with the sample application pen point 3; the filling pen core 4 is positioned in the spotting pen head 3.

The interior of the liquid suction head 1 is hollow; the liquid suction head is an elastic liquid suction head, can generate elastic deformation in an extrusion state, and can recover the original shape after the extrusion force disappears. The liquid suction head 1 is made of more than one of latex, silica gel and plastic.

Negative pressure is easily formed in the closed liquid suction head, so that liquid to be spotted cannot contact with the filling pen core, and spotting cannot be finished. Therefore, in the embodiment 1 of the present invention, the liquid suction head 1 is provided with an opening, and the opening is provided with a plug (air valve plug) with a matched size. According to the embodiment 1 of the invention, the internal and external air pressures of the liquid suction head can be adjusted by opening or closing the air valve plug, so that the liquid to be spotted can be soaked into the filling pen core, and the liquid to be spotted can be prevented from directly dripping out of the filling pen core.

The open pore can play a role in balancing air pressure, and after the liquid suction head sucks liquid, the plug can be opened to balance the air pressure inside and outside the liquid suction head, so that the sample liquid can penetrate into the sample dispensing pen core. Then the filling pen core is applied to the position needing sample application, and the sample application is completed.

The invention can adjust the internal and external air pressure of the pipette head and the liquid storage cavity by closing or opening the plug of the opening on the pipette head, thereby keeping or releasing the sample solution in the liquid storage cavity.

When the viscosity of the liquid to be spotted is small or the density is large, the plug can not be pulled out, the speed of the liquid to be spotted permeating into the filling pen core is not accelerated, and the spotting is directly finished. The invention can adjust the balance of the internal and external air pressures of the liquid suction head by pulling out the plug or not, further can control the speed of the liquid to be spotted penetrating into the filling pen core, and finally controls the spotting speed, so that the sample loading effect is more uniform, thereby being suitable for experiments limited by various conditions.

When the liquid suction head is not provided with a plug with a hole and a size corresponding to the hole, the liquid suction head can be extruded to enable the liquid in the liquid storage cavity to be in contact with the filling pen core, the liquid to be subjected to sample application is soaked into the filling pen core, and the liquid is applied to a position needing sample application through the filling pen core to complete sample application.

The opening may be provided at the top of the pipette head 1.

And a protective jacket is arranged on the periphery of the liquid storage cavity 2.

The material of the liquid storage cavity 2 is one of plastic, glass, latex, silica gel and rubber; the material of the spotting pen point 3 is more than one of plastic, glass, latex, silica gel and rubber.

The top of the sample application pen point 3 is provided with a connector which is matched with the connector at the bottom of the liquid storage cavity 2, and the connector at the bottom of the liquid storage cavity 2 is inserted into the top connector of the sample application pen point and is hermetically connected with the sample application pen point 3.

The bottom of the sample application pen point 3 is provided with an opening, and the bottom of the filling pen core 4 penetrates through the opening at the bottom of the sample application pen point 3 and extends to the outside.

When the liquid storage cavity 2 is connected with the sample application pen point 3, the top of the filling pen core 4 is tightly connected with the bottom of the liquid storage cavity 2.

The material of the refill 4 is more than one of fiber, cotton, hair and silk thread.

The volume of the liquid storage cavity is 3mL, but the volume is not limited to the volume.

The sample application pen point is the toper, the volume of sample application pen point is 200 microliters, the top diameter of sample application pen point is 6mm, the length of sample application pen point from nib to top is 50mm, but the sample application pen point is not only limited to above specification.

Example 2

Use of the spotting pen for preparing thin-layer chromatography of example 1 in thin-layer chromatography spotting, comprising the steps of:

(1) sampling: closing an air valve plug on the liquid suction head 1, extruding the liquid suction head 1 to enable the liquid suction head 1 to generate elastic deformation, inserting the bottom of the liquid storage cavity 2 into the liquid to be spotted, stopping extruding the liquid suction head to enable the liquid suction head 1 to recover the original shape, and forming negative pressure in the liquid suction head, so that the liquid to be spotted is sucked into the liquid storage cavity 2 by utilizing atmospheric pressure;

(2) connecting and loading: connecting the liquid storage cavity 2 for absorbing the liquid to be spotted in the step (1) with the spotting pen point 3, placing the spotting pen point 3 on a position needing spotting, contacting the bottom of a filling pen core 4 in the spotting pen point with the position needing spotting on the thin layer preparation plate, opening an air valve plug on the liquid absorption head 1, balancing the internal and external air pressure of the liquid absorption head 1 and the liquid storage cavity 2, and thus soaking the liquid to be spotted in the filling pen core 4; finally, the sample is applied to the position needing sample application to finish the sample application;

(3) chromatography: and (4) when the sample application liquid is volatilized, performing preparative thin-layer chromatography.

The embodiment of the invention provides an application of a thin-layer chromatography sample application pen, and the thin-layer chromatography sample application pen comprises: a liquid suction head 1 with an air valve, a liquid storage cavity 2, a point sample pen head 3 and a filling pen core 4. The liquid suction head 1 with the air valve is hermetically connected with the liquid storage cavity 2, the liquid suction head 1 is squeezed to suck sample liquid, the sample liquid is stored in the liquid storage cavity 2, the bottom of the liquid storage cavity 2 is connected to the sample application pen point 3, a plug at the top of the liquid suction head 1 with the air valve is opened to balance the internal and external air pressures, the filling pen core 4 can adsorb the sample liquid, and the sample liquid can be quickly and uniformly applied to the prepared thin-layer plate by utilizing the slight contact between the bottom of the filling pen core of the sample application pen point and the prepared thin-layer plate.

Example 3

By comparing the sample application time, sample application times, sample liquid suction times and damage number to the surface of the prepared thin layer of the thin layer chromatography sample application pen and the capillary, the thin layer chromatography sample application pen of the embodiment of the invention can obviously shorten the sample application time, greatly reduce the sample application times and the sample liquid suction times, and also reduce the damage degree to the prepared thin layer, and the specific implementation mode is as follows:

1. preparing sample liquid

Adding a sample (ferulic acid is selected) into dichloromethane, wherein the mass-volume ratio of the sample to the dichloromethane is 5: 0.4mg/mL, and after sufficient dissolution, a sample solution was obtained, and 2 portions of the sample solution, 0.1mL each, were taken, one portion for capillary loading and the other portion for loading on the sample applicator pen for thin layer chromatography preparation of example 1.

2. Preparation of the thin layer plate

Preparation of the thin-layer plate, selecting a Qingdao ocean chemical preparation thin-layer plate (the thickness is 0.4-0.5mm, the GF254 type, the length is multiplied by the width is 200mm multiplied by 100mm), drawing a parallel line with a pencil at a position 10mm away from the bottom of a long side as a sample dispensing line, marking the sample dispensing line at the midpoint, loading the sample on the left side of the midpoint by using a capillary, and loading the sample on the right side of the midpoint by using a preparation thin-layer chromatography sample dispensing pen.

3. Sample preparation and loading device

The sample was taken up by sucking up the sample liquid using a capillary (diameter 0.9-1.1mm, length 100mm) and the sample application pen of preparative thin layer chromatography of example 1, respectively.

4. Sample loading

Loading using the spotting pen of preparative thin layer chromatography of example 1: closing an air valve plug of the liquid suction head 1, extruding the liquid suction head to enable the liquid suction head 1 to generate elastic deformation, inserting the bottom of the liquid storage cavity 2 into the liquid to be spotted, stopping extruding the liquid suction head 1 to enable the liquid suction head to recover the original shape, and enabling the liquid storage cavity 2 to absorb the liquid to be spotted; and (3) hermetically connecting the liquid storage cavity 2 for absorbing the sample liquid with the sample application pen point 3, opening an air valve plug at the top of the liquid suction head to balance the internal and external air pressures, and uniformly applying the sample liquid to the prepared thin-layer plate by slightly contacting the bottom of the filling pen core 4 with a sample application standard line after the sample application liquid is soaked into the filling pen core 4.

Capillary loading: and absorbing the liquid to be spotted through the siphoning action of the capillary, sequentially loading the liquid from left to right along the spotting line, and repeating the operation until spotting is finished.

The above experiments of spotting the thin layer chromatography by the spotting pen and the capillary were repeated 3 times each, and the loading duration, spotting times, sample liquid sucking times, and number of damages to the surface of the prepared thin layer of the capillary and the spotting pen of example 1 were recorded, respectively. The results are shown in table 1 below and fig. 2.

Table 1 shows the data of the loading time, the number of spotting times, the number of times of sucking the sample liquid, and the number of damages to the surface of the prepared sheet for the capillary and the spotting pen of example 1. Part a of fig. 2 is a graph showing surface damage of a thin layer after loading by a capillary, and part b of fig. 2 is a graph showing surface damage of a thin layer after loading by the application pen for preparative thin layer chromatography.

TABLE 1

As can be seen from table 1 and fig. 2, the spotting pen of the embodiment of the present invention can take up the sample solution at one time and complete the spotting 1 time under the same loading volume. However, the application of the traditional capillary sample loading not only has more times of absorbing sample liquid and correspondingly increases the number of sample application, but also has long total sample application time and more corresponding thin layer surface damage; when the sample application pen for preparing the thin-layer chromatography provided by the embodiment of the invention is used for loading, the number of damaged surfaces of thin layers and the loading time are obviously reduced.

Example 4

Compared with the sample loading time and the sample loading effect of a thin-layer chromatography sample application pen and a capillary, the thin-layer chromatography sample application pen provided by the embodiment of the invention obviously shortens the sample application time and can make the sample application spots more uniform, and the specific implementation mode is as follows:

1. preparation of sample solution

A. Preparing a fat-soluble sample liquid: adding a sample (ferulic acid is selected) into dichloromethane, wherein the mass-volume ratio of the sample to the dichloromethane is 5: 0.4mg/mL, and after sufficiently dissolving to obtain a sample solution, 2 parts of the sample solution (liposoluble sample solution) were added by 0.1mL, one part was used for sample application by a sample application pen for preparative thin layer chromatography in example 1, and the other part was used for capillary sample application.

B. Preparation of water-soluble sample solution: adding a sample (caffeine is selected) into water, wherein the mass volume ratio of the sample to dichloromethane is 5: 0.4mg/mL, and after sufficiently dissolving to obtain a sample solution, 2 parts of the sample solution (water-soluble sample solution) were added by 0.1mL, one part was used for sample application by a sample application pen for preparative thin layer chromatography in example 1, and the other part was used for capillary application. .

2. Preparation of the thin layer plate

Preparing the thin-layer plate, selecting a Qingdao ocean chemical preparation thin-layer plate (the thickness is 0.4-0.5mm, the GF254 type, the length is multiplied by the width is 200mm multiplied by 100mm), using a pencil as a parallel line which is 10mm away from the bottom of a long side as a standard line for sample application, marking the midpoint of the standard line of the long side, applying samples on the left side of the thin-layer plate by using a capillary, and applying samples on the right side of the thin-layer plate by using a sample application pen.

3. Sample preparation and loading device

The sample was loaded by sucking the sample solution using the sample application pen and capillary (diameter 0.9mm to 1.1mm, length 100mm) for thin layer chromatography of example 1, respectively.

4. Sample loading

Sample application pen for preparation of thin layer chromatography of example 1: closing an air valve plug of the liquid suction head 1, extruding the liquid suction head to enable the liquid suction head 1 to generate elastic deformation, inserting the bottom of the liquid storage cavity 2 into the liquid to be spotted, stopping extruding the liquid suction head to enable the liquid suction head 1 to recover the original shape, and enabling the liquid storage cavity 2 to absorb the liquid to be spotted; the liquid storage cavity 2 for absorbing the sample liquid is hermetically connected with the sample application pen point 3, the air valve plug at the top of the liquid absorption head 1 is opened to balance the internal and external air pressure, the sample application liquid is soaked into the filling pen core 4, and the bottom of the filling pen core 4 slightly contacts with the sample application standard line to uniformly apply the sample liquid to the prepared thin-layer plate.

Capillary loading: and absorbing the liquid to be spotted through the siphoning action of the capillary, sequentially loading the liquid from left to right along the spotting line, and repeating the operation until spotting is finished.

The loading time (table 2) and loading effect (see figure 3) of the capillary and the applicator pen of example 1 were recorded separately. Table 2 shows the loading times of the capillary and the spotter pen of example 1. FIG. 3 is a graph showing the effect of the capillary and the applicator pen for preparing thin layer chromatography in example 4 of the present invention after loading.

Part a1 and part a2 in fig. 3 are graphs of the effect of capillary loading, wherein part a1 is a fat-soluble sample and part a2 is a water-soluble sample; part b1 and part b2 in fig. 3 are graphs of the effect of loading the sample application pen, wherein part b1 is a fat-soluble sample and part b2 is a water-soluble sample. As can be seen from fig. 3, when the capillary is used for loading, the repeated application of the fat-soluble sample for many times often causes uneven overlapping arrangement of the dotted spots, and the water-soluble sample liquid causes the problem of too wide spread of the loading color band.

TABLE 2

As can be seen from table 2 and fig. 3, the thin layer chromatography spotting pen of the embodiment of the present invention significantly shortens the spotting time, does not spread too wide the sample band, and can make the distribution of the sample applied to the spotting line more uniform, compared to the capillary spotting.

The above examples are only preferred embodiments of the present invention, which are intended to be illustrative and not limiting, and those skilled in the art should understand that they can make various changes, substitutions and alterations without departing from the spirit and scope of the invention.