Rapid identification method of ocimum gratissimum

Abstract

The invention relates to the technical field of plant analysis and detection, in particular to a rapid identification method of ocimum gratissimum. From a brand new perspective, through comparing the morphological characteristics of the ocimum basilicum seed colloid and the starch particles in the colloid tube, the ocimum gratissimum can be quickly and accurately distinguished from other ocimum species by utilizing the morphological characteristics of the ocimum basilicum seed colloid and the starch particles in the colloid tube, so that the ocimum gratissimum and other ocimum species can be quickly and accurately distinguished. The method has important significance on application and industrial development of the ocimum gratissimum.

Description

Technical Field

[0001] The present invention relates to the technical field of plant analysis and detection, and particularly relates to a rapid identification method for Ocimum gratissimum. Background Art

[0002] Ocimum belongs to the Lamiaceae family, including annual or perennial herbs or shrubs. As an aromatic medicinal plant, it has been widely recognized by people all over the world due to its extensive applications in medicine, food, and daily chemical products. Currently, the commonly used Ocimum species mainly include Ocimum basilicum, Ocimum gratissimum, Ocimum sanctum, etc., and they are widely used in the treatment of gastrointestinal diseases, eye diseases, colds and coughs, joint diseases, pain relief, etc. Moreover, Ocimum is also used to treat skin diseases, respiratory diseases, urinary diseases, and diabetes; in addition, some people use it to repel mosquitoes. Due to the extensive interspecific and intraspecific hybridization, polyploidy, aneuploidy, various local varieties and cultivated varieties in Ocimum, Ocimum has rich genetic diversity of resources. Pushpangadan and Bradu identified 160 Ocimum species in 1995, while Paton et al. confirmed 64 Ocimum species in the overview of the classification and relationships of Ocimum in 1999. And Ocimum has a wide distribution range and a long application history, thus forming different names around the world. However, this has led to the widespread existence of the situations of different names for the same plant and the same name for different plants, which is very easy to cause confusion and brings a series of difficulties to the origin identification of plant medicinal materials of this genus, the formulation of quality standards, and the application, etc.

[0003] Ocimum gratissimum is an important species in the genus Ocimum, having multiple subspecies, such as the original species of Ocimum gratissimum, Ocimum gratissimum var. suave, Ocimum gratissimum var. macroghyllum, etc. In the early stage, our research group collected 155 Ocimum resources from more than 30 countries and regions and conducted botanical investigations. It was found that there were significant morphological differences among different Ocimum gratissimum resources. The leaf sizes varied from 2 to 10 cm. Some leaves, leaf petioles, and stems were covered with villi, while in some Ocimum gratissimum germplasms with special origins, the villi on the leaves and leaf petioles were relatively scarce, and the floral structure characteristics also differed greatly. Generally, they were white, while in some materials from certain origins, purple patterns were shown on the petals. The leaf shapes of Ocimum gratissimum var. microphyllum and Ocimum sanctum are extremely similar and difficult to distinguish ( Figure 1 ).

[0004] Therefore, how to provide a method to quickly and accurately distinguish Ocimum gratissimum from other species in the genus Ocimum has become a technical problem urgently to be solved in this field. Summary of the Invention

[0005] To solve the above technical problems, through a large number of comparisons of the gelation states of the seed colloids of Ocimum gratissimum and other Ocimum species in distilled water, it is found that there are obvious differences in the gelation states of the seed colloids between Ocimum gratissimum and other Ocimum species, especially the length of the seed colloid and the size of the starch granules in its colloidal tube. Based on this, the following technical solutions are proposed.

[0006] In a first aspect, the present invention provides an application of Ocimum seed colloid in identifying Ocimum gratissimum.

[0007] In a specific implementation process, the Ocimum gratissimum is identified by using the length of the Ocimum seed colloid and the size of the starch granules in its colloidal tube.

[0008] In a second aspect, the present invention provides an application of a reagent for detecting the Ocimum seed colloid and the starch granules in its colloidal tube in the preparation of a product for identifying Ocimum gratissimum.

[0009] In a specific implementation process, the product includes but is not limited to a detection reagent or a detection kit, etc.

[0010] In some embodiments, the reagent is used to detect the length of the Ocimum seed colloid, the size of the starch granules in the colloidal tube, the loose state of the colloidal tube, or the aggregation of the starch granules.

[0011] In a third aspect, the present invention provides a rapid identification method for Ocimum gratissimum, including: judging whether the test Ocimum is Ocimum gratissimum by observing the gelation state of the seed colloid in distilled water in the seed sample of the test Ocimum.

[0012] In a specific implementation process, if the colloid length is 0.1 - 0.2 cm and the size of a single granule in the colloidal tube is 5 - 15 μm, then it is judged that the test Ocimum is Ocimum gratissimum.

[0013] In a specific implementation process, compared with the seed samples of other test Ocimum, if the granules in the colloidal tube are in an aggregated form of aggregation and / or the colloidal tube is in a loose state, then it is further confirmed that the test Ocimum is Ocimum gratissimum.

[0014] Furthermore, the method further includes: further verifying whether the test Ocimum is Ocimum gratissimum according to the staining condition of the granules in the colloidal tube. When the test Ocimum is Ocimum gratissimum, the granules in the colloidal tube can be dyed purple by an iodophor solution, indicating that the granules are starch granules.

[0015] Furthermore, the method further includes: further verifying whether the test Ocimum is Ocimum gratissimum according to the granule morphology in the colloidal tube under a polarized light microscope. When the test Ocimum is Ocimum gratissimum, the granule morphology has more than 2 umbilici, and the granules are compound grains.

[0016] Preferably, the povidone iodine solution is a 10% povidone iodine solution.

[0017] In the specific implementation process, the method further includes: further verifying whether the basil to be tested is Ocimum gratissimum according to the particle staining situation in the colloidal tube. When the basil to be tested is Ocimum gratissimum, the seed sample is spherical or nearly spherical, reddish-brown, and has dark brown patterns on the surface.

[0018] In the specific implementation process, a stereoscopic fluorescence microscope or a polarized light microscope is used for observation.

[0019] Preferably, a DS-Ri2 digital imaging system is adopted, and after shooting and synthesizing an image through depth of field extension, observation is carried out.

[0020] Compared with the prior art, the beneficial effects of the present invention are as follows:

[0021] From a brand-new perspective, the present invention discovers that by comparing the morphological characteristics of the starch particles in the basil seed colloid and its colloidal tube, Ocimum gratissimum can be quickly and accurately distinguished from other Ocimum species by using the morphological characteristics of the starch particles in the basil seed colloid and its colloidal tube. Therefore, the present invention is of great significance for the application and industrial development of Ocimum gratissimum. Description of the Drawings

[0022] Figure 1 It is a diagram of the leaves of different germplasm Ocimum gratissimum (G144, G134, G096) and the easily confused Ocimum tenuiflorum leaves (G060).

[0023] Figure 2 It is a distribution diagram of the colloidal properties of Ocimum gratissimum G131 seeds and the particles in the colloidal tube. Among them, A is a photo of the seeds; B-D are photos of the colloidal tube and the particles in the colloidal tube; E and F are diagrams of the staining results.

[0024] Figure 3 It is a distribution diagram of the colloidal properties of Ocimum gratissimum G122, G141, G145 seeds and the particles in the colloidal tube. Among them, A is a photo of G122 seeds; B is a photo of the G122 colloidal tube and the particles in the colloidal tube; C is a photo of G141 seeds; D is a photo of the G141 colloidal tube and the particles in the colloidal tube; E is a photo of G145 seeds; F is a photo of the G145 colloidal tube and the particles in the colloidal tube.

[0025] Figure 4It is a distribution diagram of the colloidal properties of the seeds of other species of the Ocimum genus and the particles in the colloidal tubes. Among them, A is a photo of the seeds of Ocimum basilicum G002; B is a photo of the colloidal tube of Ocimum basilicum G002; C is a stained result diagram of the particles in the colloidal tube of Ocimum basilicum G002; D is a photo of the seeds of Ocimum sanctum G058; E is a photo of the colloidal tube of Ocimum sanctum G058; F is a stained result diagram of the particles in the colloidal tube of Ocimum sanctum G058; G is a photo of the seeds of Ocimum gratissimum G095; H is a photo of the colloidal tube of Ocimum gratissimum G095; I is a stained result diagram of the particles in the colloidal tube of Ocimum gratissimum G095; J is a photo of the seeds of Ocimum × Africnum G066; K is a photo of the colloidal tube of Ocimum × Africnum G066; L is a stained result diagram of the particles in the colloidal tube of Ocimum × Africnum G066.

[0026] Figure 5 It is a photo of the starch granules in the colloidal tube of Ocimum gratissimum G141 under a polarized light microscope. Detailed implementation manners

[0027] To make the objectives, technical solutions and advantages of the present invention clearer, the technical solutions in the present invention will be clearly and completely described below. Apparently, the described embodiments are some but not all of the embodiments of the present invention. All other embodiments obtained by those of ordinary skill in the art without making creative efforts based on the embodiments in the present invention belong to the scope of protection of the present invention.

[0028] In the embodiments provided in this specification, for those without specific technical or conditions indicated, the technologies or conditions described in the literature in the field are followed, or the product specifications are followed. For the reagents or instruments without the manufacturer indicated, they are all conventional products that can be obtained through regular channels.

[0029] Embodiment

[0030] 1. Select 5 - 10 dry seeds of different species resources of the Ocimum genus, place them on the stage of a stereoscopic fluorescence microscope (Leica M205FA), and place them naturally so that the picture shows a natural state of not being crowded and loose. Use the DS-Ri2 digital imaging system to take and synthesize images by depth of field extension. Compare the morphological differences of the dry seeds of different species of the Ocimum genus. It can be seen that the seeds of Ocimum gratissimum are generally nearly spherical, reddish-brown, and have dark brown patterns on the surface ( Figure 2 A, Figure 3 A, Figure 3 C, Figure 3 E). While the seeds of other species of Ocimum such as Ocimum basilicum and Ocimum × Africnum G066 are nearly black in color and have reticulate protrusions on the surface ( Figure 4 A, Figure 4J). The seeds of Ocimum campechianum are reddish-brown and relatively flat. Figure 4 G); while the seeds of Ocimum sanctum are slightly similar to those of Ocimum gratissimum, being spherical as a whole, reddish-brown or black. Figure 4 D). The seed sizes are in the order of Ocimum basilicum > Ocimum campechianum > Ocimum gratissimum > Ocimum sanctum > Ocimum × africanum.

[0031] 2. Select representative seeds, place them on a glass slide, and drop distilled water on the seed surface. Also use the above method for depth of field extension and take synthetic images to observe the gelling state of the basil seed colloids of different species in the genus Ocimum in distilled water.

[0032] It can be seen that except for Ocimum gratissimum, white colloids like fluff are emitted from the surfaces of the seeds of different species in the genus Ocimum, with a length of 0.25 - 0.5 cm; upon careful observation, it can be seen that the colloids are tubular, and white particles are evenly dispersed in the tubular bodies of the basil seed colloids, extending from the seed coat surface to the top of the colloid tube. The size of the middle white particles and the number scattered in each colloid tube vary with the species. Figure 4 B, Figure 4 E, Figure 4 H, Figure 4 K). In Ocimum gratissimum, the length of the colloid is shorter, only 0.1 - 0.2 cm, and the colloid tubes are looser. It should be noted that the particles in the colloid tubes of Ocimum gratissimum are all aggregated together in a clustered form, and the individual particles themselves are larger than those of other species and are very easy to identify.

[0033] To further confirm whether other subspecies or germplasms from other sources of Ocimum gratissimum also have this characteristic, other resources such as Ocimum gratissimum G122 from Madagascar, Ocimum gratissimum G141 from Thailand, and Ocimum gratissimum G145 from Ethiopia were additionally selected for observation. The results all showed that the seeds are nearly spherical, reddish-brown, with dark brown patterns on the surface, the colloid tubes are looser and the length of the colloid is shorter, and the particles in the colloid tubes are larger and in a clustered form. Figure 3 B, Figure 3 D, Figure 3 F), and it is extremely easy to identify the Ocimum gratissimum species from other species in the genus Ocimum. Only as an example, the photo of the starch particles in the colloid tube of Ocimum gratissimum G141 under a polarized light microscope is as Figure 5 shown. To ensure the accuracy of the experiment, the present invention also tested 6 basil species of other 32 germplasms such as O. basilicum, O. × Africnum, O. campechianum, O. americanum, O. kilimandscharicum, O. tenuiflorum, etc., and all showed the particularity of the particle morphology and distribution in the Ocimum gratissimum seed colloids. Some test results are shown in Table 1.

[0034] Table 1

[0035]

[0036]

[0037] 3. Further add 10% povidone iodine solution to the basil seeds soaked in distilled water. The results show that the particles in the colloids of the seeds of all Ocimum species can be dyed purple, indicating that the particles in the basil seed colloids are starch particles. Figure 2 E, Figure 2 F, Figure 4 C, Figure 4 F, Figure 4 I, Figure 4 L).

[0038] Finally, it should be noted that the above embodiments are only used to illustrate the technical solutions of the present invention, rather than to limit it; although the present invention has been described in detail with reference to the foregoing embodiments, those of ordinary skill in the art should understand that they can still modify the technical solutions recorded in the foregoing embodiments, or perform equivalent replacements on some of the technical features; and these modifications or replacements do not make the essence of the corresponding technical solutions deviate from the spirit and scope of the technical solutions of the embodiments of the present invention.

Claims

1. Application of basil seed colloid and starch granules in its colloid tube in identifying basil.

2. Application of reagents for detecting basil seed colloids and starch granules in basil seed colloid tubes in the preparation of products for identifying basil.

3. A rapid identification method for basil, characterized in that: include: Whether the basil to be tested is clove basil can be determined by observing the gelling state of the seed colloid in the seed sample of the basil to be tested in distilled water.

4. The method according to claim 3, characterized in that If the length of the colloid is 0.1-0.2 cm, and the size of a single particle in the colloid tube is 5-15 um, the basil to be tested is determined to be clove basil.

5. The method according to claim 4, characterized in that Compared with other test basil seed samples, if the particles in the colloid tube are in a clustered aggregated form, and / or the colloid tube is in a loose state, it is further confirmed that the basil to be tested is clove basil.

6. The method according to any one of claims 3 to 5, characterized in that: Also includes: Whether the basil to be tested is clove basil is further verified based on the staining of the particles in the colloid tube. When the basil to be tested is clove basil, the particles in the colloid tube can be dyed purple by iodine solution, indicating that the particles are starch particles.

7. The method according to claim 6, characterized in that The starch granule morphology in the colloid tube under a polarizing microscope was used to further verify whether the basil to be tested was Eugenia sylvestris. When the basil to be tested was Eugenia sylvestris, the starch granule morphology had more than two hilums and the starch granules were complex grains.

8. The method according to any one of claims 3 to 5, characterized in that: Also includes: Whether the basil to be tested is clove basil is further verified based on the staining of the particles in the colloid tube. When the basil to be tested is clove basil, the seed sample is spherical or nearly spherical, reddish brown, and has dark brown patterns on the surface.

9. The method according to any one of claims 3 to 5, characterized in that: Observe using a fluorescent stereo microscope or a polarizing microscope.

10. The method according to claim 9, characterized in that The DS-Ri2 digital imaging system was used to take composite images with extended depth of field for observation.

Images