CN105199425A - Extraction method of cyanidin - Google Patents
Extraction method of cyanidin Download PDFInfo
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- CN105199425A CN105199425A CN201510655494.1A CN201510655494A CN105199425A CN 105199425 A CN105199425 A CN 105199425A CN 201510655494 A CN201510655494 A CN 201510655494A CN 105199425 A CN105199425 A CN 105199425A
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Abstract
The invention belongs to the field of plant effective component extraction, and provides an extraction method of cyanidin. The method comprises the following steps: (1) sample preparation; (2) leaf grinding; and (3) cyanidin extraction. The orthogonal test by using a citric acid-sodium citrate buffer solution as an extraction solvent detects that the sequence of factors responding to the purple lettuce cyanidin extraction effect is as follows: material/liquid ratio>extraction temperature>extraction time; and therefore, the material/liquid ratio is preferentially determined in the cyanidin extraction process. The control on the material/liquid ratio, extraction time, extraction temperature and extracting solution pH value proves that the method enhances the extraction rate and cyanidin leaching rate on the premise of lowering the cost and simplifying the process. Since all the extraction solvents are nontoxic and are reagents widely used as food additives, the method can satisfy the requirements of modern people for safety, and thus, has wide applicability in the fields of food, health care, drugs and the like.
Description
Technical field
The invention belongs to effective ingredients in plant and extract field, relate to a kind of extracting method of anthocyanidin.
Background technology
Anthocyanidin (Anthocyanin), also known as cyanidin(e), is that a class is extensively present in natural pigment in plant, and belong to flavonoid compound, the form mainly with glucosides exists, and is the main colour former of plant.Anthocyanidin is as natural pigment; there is higher safety performance; and there is anti-mutation, hypotensive, nourishing function such as protection liver etc.; and anthocyanidin is in Europe; be called as " oral skin cosmetics ", can prevent generating ahead of time of wrinkle of skin, be the most effective antioxidant of current nature; not only can prevent generating ahead of time of wrinkle of skin, more can supplement the nutrients and eliminate free radical harmful in body.Modern finds, although the research of microbiotic and VITAMIN is very deep, also can't resolve modern diseases and the sub-health status such as such as cardiovascular and cerebrovascular diseases, diabetes, cancer, more can not solve the promoting longevity of people, antidotal problem.Scientific research: if once solve the infringement problem of free radical, so human body cell just can really freely be grown up, and the mean lifetime of people is bound to reach 125 years old.So the life-span length of people directly depends on the power of the anti-oxidant antiradical activities of people, and anthocyanidin be found to be the most simple and effective way that global people have found antioxidant and anti-aging.The discovery of anthocyanidin and application make the mankind from the microbiotic in 20th century, VITAMIN epoch, enter into the anthocyanidin epoch of 21 century! Along with the development of science and technology and the raising of living standards of the people, the security of people to foodstuff additive and natural pigment series products is more and more paid attention to, the use kind of synthetic colour and quantity significantly decline, therefore, development and application natural pigment has become the general trend of world's food dye development.
Anthocyanidin is water colo(u)r, soluble in water, the alcohol compound such as ethanol and methyl alcohol.Anthocyanidin is unstable in neutrality and weak basic condition.Therefore, leaching process adopts acid solvent usually.The selection of Extraction medium is an important link in nature strength process, applies minimum subsidiary and reaches maximum pigment extracted amount with the minimal degradation of target product or change.In existing report, methyl alcohol, acetone, hydrochloric acid etc. can be used for extracting anthocyanin, but due to the genotoxic potential of residual solvent, and extract is dangerous for human consumer.So, for the selection of anthocyanidin Extraction solvent, should select food can solvent.
Summary of the invention
The object of the invention is for existing issue, a kind of extracting method of anthocyanidin is provided.
Object of the present invention is achieved through the following technical solutions
1. the invention provides a kind of extracting method of anthocyanidin, the method comprises the steps:
(1) sample prepares: get fresh blade sample, cleans, dries, and removes main lobe arteries and veins, shreds blade and weigh, be placed in mortar, stand-by;
(2) blade grinding: according to solid-liquid ratio 1: 5g/ml, add the citric acid-sodium citrate buffer that pH is 2-4 in mortar; Under condition of ice bath, grinding pulping; Feed liquid proceeds to centrifuge tube in the lump; Continue in mortar, add the citric acid-sodium citrate buffer that pH is 2-4, after rinsing mortar, proceed to above-mentioned centrifuge tube, make final solid-liquid ratio be 1: 50-1: 10;
(3) anthocyanidin extracts: be centrifuge tube lucifuge concussion 20-100min extraction anthocyanidin at 30-60 DEG C of 1: 50-1: 10 by final solid-liquid ratio, centrifugal; Supernatant liquor is anthocyanidin solution after filter paper suction filtration.
2. the extracting method of above-mentioned 1 anthocyanidin provided, wherein, the final solid-liquid ratio described in step (2) is 1: 30.
3. the extracting method of above-mentioned 1 anthocyanidin provided, wherein, the pH of the citric acid-sodium citrate buffer that step (2) is used is 3.
4. the extracting method of above-mentioned 1 anthocyanidin provided, wherein, the Extracting temperature described in step (2) is 50 DEG C.
5. the extracting method of above-mentioned 1 anthocyanidin provided, wherein, the extraction time described in step (2) is 60min.
6. above-mentioned any one of 1-5 provides the extracting method of anthocyanidin, and wherein, step (3) centrifugal condition is 4 DEG C, 4000rpm/min, centrifugal 10min.
7. the application of above-mentioned any one of 1-6 extracting method in purple romaine lettuce, purple cabbage in anthocyanidin extraction.
8. the anthocyanidin solution that the extracting method provided according to above-mentioned any one of 1-6 obtains.
9. the above-mentioned 8 anthocyanidin solution provided, wherein, in anthocyanidin solution, relative anthocyanidin content U is 0.071.
Beneficial effect:
1. the present invention adopts food useable solvents citric acid-sodium citrate buffer as the Extraction solvent of anthocyanidin, do not produce pollution, anthocyanidin after extraction is without poisonous and harmful solvent, safe and reliable, can meet the requirement of modern to safety, thus it has a wide range of applications in fields such as food, health care, medicines.
2. the present invention is using citric acid-sodium citrate buffer as extracting solution, and by optimizing leaching process, finally, in the anthocyanidin solution of generation, relative anthocyanidin content U is 0.071.When reaching optimum condition, namely citric acid-sodium citrate buffer pH is 3, and solid-liquid ratio is 1: 30, Extracting temperature 50 DEG C, extraction time 60min, relative anthocyanidin content U reaches 0.071, be 0.038 relative to the relative anthocyanidin content U of traditional method for extracting, be significantly improved.
3. the present invention does not adopt complicated step and solvent, is reducing costs, and while simplifying extraction step, improves the solubility rate of anthocyanidin, can reach 86.8%.
Accompanying drawing explanation
The impact of Fig. 1 different feed liquid comparison anthocyanidin content
Fig. 2 different extraction time is on the impact of anthocyanidin content
The different Extracting temperature of Fig. 3 is on the impact of anthocyanidin content
The different extracting liquid pH value of Fig. 4 is on the impact of anthocyanidin content
Embodiment
Below in conjunction with embodiment, the present invention will be further described, and the experimental technique of unreceipted actual conditions in the following example, usually according to the known approaches of this area.
Embodiment 1
1. design solid-liquid ratio to be respectively 1: 20,1: 30,1: 40 (m/v, g/ml), Extracting temperature 40,50,60 DEG C, each extraction time 40,60,80min.Design orthogonal test, level of factor is see table 1.
2. get fresh blade, clean, dry, remove main lobe arteries and veins, shred blade and be weighed as 0.4g, be placed in mortar.First adding a little in mortar is the citric acid-sodium citrate buffer of 3 through the pH of precooling, after grinding homogenate, proceeds in centrifuge tube under condition of ice bath.
Table 1 quadrature factor water-glass
3. according to the orthogonal test that above-mentioned steps 1 designs, in mortar, repeatedly add citric acid-sodium citrate buffer respectively rinse mortar, washing fluid is proceeded to centrifuge tube, mixing, final solid-liquid ratio is made to be respectively 1: 20,1: 30,1: 40 (m/v, g/ml), the centrifuge tube of each solid-liquid ratio respectively at 40-60 DEG C lucifuge concussion extract 40-80min, carry out orthogonal test in; Afterwards in 4 DEG C, 4000rpm/min, centrifugal 10min.Supernatant liquor is sample solution after filter paper suction filtration.
4. Specimen Determination: sample solution extracting solution is settled to 100ml, ultraviolet spectrophotometer is utilized to measure absorbancy respectively at wavelength 530nm and 600nm place, anthocyanidin content (U) is represented, i.e. U=(OD530-OD600)/g by the difference of 530nm and 600nm place absorbancy.Result is as table 2, can be obtained by table 2, the response primary and secondary order of each factor to purple romaine lettuce anthocyanidin extraction effect is: A (solid-liquid ratio) > B (Extracting temperature) > C (extraction time), therefore first should determine solid-liquid ratio in purple romaine lettuce anthocyanidin extraction process.
Table 2 orthogonal experiments
K1, k2, k3, expression average; R represents that extreme difference in general, and extreme difference is larger, and the impact of this factor on index is larger.
Embodiment 2
1. get fresh blade and amount to 15 parts, clean, dry, remove main lobe arteries and veins, shred blade and weigh, every part is 0.4g, is placed on respectively in mortar.
2. be the citric acid-sodium citrate buffer of 3 respectively to adding a little in mortar through the pH of precooling, grind homogenate under condition of ice bath after, feed liquid proceeds in centrifuge tube in the lump, numbering.Continue respectively to add citric acid-sodium citrate buffer in mortar, repeatedly rinse mortar, proceed in the centrifuge tube of above-mentioned correspondence afterwards respectively, mixing, finally, solid-liquid ratio is 1: 10,1: 20,1: 30,1: 40,1: 50 (m/v, g/ml), and each feed liquid ratio repeats for 3 times, feed liquid lucifuge concussion extraction 60min at 50 DEG C respectively in centrifuge tube, in 4 DEG C, 4000rpm/min, centrifugal 10min.Supernatant liquor is sample solution after filter paper suction filtration.
3. Specimen Determination: sample solution extracting solution is settled to 100ml, ultraviolet spectrophotometer is utilized to measure absorbancy respectively at wavelength 530nm and 600nm place, anthocyanidin content (U) is represented, i.e. U=(OD530-OD600)/g by the difference of 530nm and 600nm place absorbancy.Result is see Fig. 1.Can obtain by Fig. 1, when solid-liquid ratio is 1: 30, purple leaf romaine lettuce anthocyanidin extracted amount is maximum, reaches conspicuous level.
Embodiment 3
1. get fresh blade and amount to 15 parts, clean, dry, remove main lobe arteries and veins, shred blade and weigh, every part of 0.4g, be placed on respectively in mortar.
2. be the citric acid-sodium citrate buffer of 3 respectively to first adding a little in mortar through the pH of precooling, grind homogenate under condition of ice bath after, feed liquid proceeds in centrifuge tube in the lump, numbering.Continue respectively to add citric acid-sodium citrate buffer in mortar, repeatedly rinse mortar, proceed in above-mentioned centrifuge tube afterwards respectively, mixing, finally, solid-liquid ratio is 1: 30 (m/v, g/ml), feed liquid lucifuge concussion extraction 20 at 50 DEG C respectively in centrifuge tube, 40,60,80,100min, each extraction time repeats 3 times, centrifuge tube in 4 DEG C, 4000rpm/min, centrifugal 10min.Supernatant liquor is sample solution after filter paper suction filtration.
3. Specimen Determination: sample solution extracting solution is settled to 100ml, ultraviolet spectrophotometer is utilized to measure absorbancy respectively at wavelength 530nm and 600nm place, anthocyanidin content (U) is represented, i.e. U=(OD530-OD600)/g by the difference of 530nm and 600nm place absorbancy.Result is see Fig. 2.Obtained by Fig. 2, when being 60min upon extracting, purple leaf romaine lettuce anthocyanidin extracted amount is maximum, reaches conspicuous level.
Embodiment 4
1. get 12 parts, fresh blade, clean, dry, remove main lobe arteries and veins, shred blade and weigh, every part of 0.4g, be placed on respectively in mortar.
2. be the citric acid-sodium citrate buffer of 3 respectively to first adding a little in mortar through the pH of precooling, grind homogenate under condition of ice bath after, feed liquid proceeds in centrifuge tube in the lump, numbering.Continue respectively to add citric acid-sodium citrate buffer in mortar, repeatedly rinse mortar, proceed in above-mentioned centrifuge tube afterwards respectively, mixing, finally, solid-liquid ratio is 1: 30 (m/v, g/ml), feed liquid lucifuge concussion extraction 60min at 30,40,50,60 DEG C respectively in centrifuge tube, each Extracting temperature repeats 3 times, centrifuge tube in 4 DEG C, 4000rpm/min, centrifugal 10min.Supernatant liquor is sample solution after filter paper suction filtration.
3. Specimen Determination: sample solution extracting solution is settled to 100ml, ultraviolet spectrophotometer is utilized to measure absorbancy respectively at wavelength 530nm and 600nm place, anthocyanidin content (U) is represented, i.e. U=(OD530-OD600)/g by the difference of 530nm and 600nm place absorbancy.Result is see Fig. 3.Can be obtained by Fig. 3, when Extracting temperature is 50 DEG C, purple leaf romaine lettuce anthocyanidin extracted amount is maximum, reaches conspicuous level.
Embodiment 5
1. get fresh blade and amount to 15 parts, clean, dry, remove main lobe arteries and veins, shred blade and weigh, every part of 0.4g, be placed on respectively in mortar.
2. respectively to first add in mortar a little through the pH of precooling be 2,2.5,3,3.5,4 citric acid-sodium citrate buffer, the buffered soln of each pH repeats 3 times, and grind homogenate under condition of ice bath after, feed liquid proceeds in centrifuge tube in the lump, numbering.Continue respectively to add citric acid-sodium citrate buffer in mortar, repeatedly rinse mortar, proceed to respectively afterwards in above-mentioned centrifuge tube, mixing, finally, solid-liquid ratio is 1: 30 (m/v, g/ml), feed liquid lucifuge concussion extraction 60min at 50 DEG C respectively in centrifuge tube, centrifuge tube is in 4 DEG C, 4000rpm/min, centrifugal 10min.Supernatant liquor is sample solution after filter paper suction filtration.
3. Specimen Determination: sample solution extracting solution is settled to 100ml, ultraviolet spectrophotometer is utilized to measure absorbancy respectively at wavelength 530nm and 600nm place, anthocyanidin content (U) is represented, i.e. U=(OD530-OD600)/g by the difference of 530nm and 600nm place absorbancy.Result is see Fig. 4.Obtained by Fig. 4, anthocyanidin is when pH is 2 ~ 3, and anthocyanidin is comparatively stable.When extracting solution pH is 3, purple leaf romaine lettuce anthocyanidin extracted amount is maximum, reaches conspicuous level.
Can know; above-described embodiment is only in order to illustrate the illustrative embodiments that inventive principle adopts; but the present invention is not limited only to this; those skilled in the art are not departing under real situation of the present invention; can make various improvement and change, these improve and change and also belong to protection scope of the present invention.
Claims (9)
1. an extracting method for anthocyanidin, is characterized in that, the method comprises the steps:
(1) sample prepares: get fresh blade sample, cleans, dries, and removes main lobe arteries and veins, shreds blade and weigh, be placed in mortar, stand-by;
(2) blade grinding: according to solid-liquid ratio 1: 5g/ml, add the citric acid-sodium citrate buffer that pH is 2-4 in mortar; Under condition of ice bath, grinding pulping; Feed liquid proceeds to centrifuge tube in the lump; Continue in mortar, add the citric acid-sodium citrate buffer that pH is 2-4, after rinsing mortar, proceed to above-mentioned centrifuge tube, make final solid-liquid ratio be 1: 50-1: 10;
(3) anthocyanidin extracts: be centrifuge tube lucifuge concussion 20-100min extraction anthocyanidin at 30-60 DEG C of 1: 50-1: 10 by final solid-liquid ratio, centrifugal; Supernatant liquor is anthocyanidin solution after filter paper suction filtration.
2. the extracting method of anthocyanidin described in claim 1, is characterized in that: the final solid-liquid ratio described in step (2) is 1: 30.
3. the extracting method of anthocyanidin described in claim 1, is characterized in that: the pH of the citric acid-sodium citrate buffer that step (2) is used is 3.
4. the extracting method of anthocyanidin described in claim 1, is characterized in that: the Extracting temperature described in step (2) is 50 DEG C.
5. the extracting method of anthocyanidin described in claim 1, is characterized in that: the extraction time described in step (2) is 60min.
6. the extracting method of anthocyanidin described in any one of claim 1-5, is characterized in that: step (3) centrifugal condition is 4 DEG C, 4000rpm/min, centrifugal 10min.
7. the application of any one of claim 1-6 extracting method in purple romaine lettuce, purple cabbage in anthocyanidin extraction.
8. the anthocyanidin solution that extracting method obtains according to any one of claim 1-6.
9. anthocyanidin solution according to claim 8, is characterized in that: in anthocyanidin solution, relative anthocyanidin content U is 0.071.
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Cited By (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN105503981A (en) * | 2015-12-01 | 2016-04-20 | 大兴安岭林格贝寒带生物科技股份有限公司 | Method for extracting and separating anthocyanin from red cabbage |
| CN109577031A (en) * | 2018-12-27 | 2019-04-05 | 河北科技大学 | Using vegetable colour to the method for textile dyeing |
| CN109588568A (en) * | 2018-12-18 | 2019-04-09 | 南京农业大学 | A method of procyanidine is added by daily ration and improves sow reproductive performance |
Citations (6)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| JPH01132344A (en) * | 1988-10-21 | 1989-05-24 | T Hasegawa Co Ltd | Food and drink |
| CN101735647A (en) * | 2009-12-29 | 2010-06-16 | 威海华彩盛业生物科技有限公司 | Method for extracting ipomoes batatas L. pigment |
| CN101735644A (en) * | 2009-11-20 | 2010-06-16 | 苏州大学 | Method for extracting ipomoes batatas L. anthocyanidins |
| CN101186757B (en) * | 2007-12-24 | 2010-10-06 | 钱生球 | Method for producing purple sweet potato haematochrome |
| CN102206426A (en) * | 2011-03-23 | 2011-10-05 | 武汉普赛特膜技术循环利用有限公司 | Comprehensive utilization of pigment of purple sweet potato in purple sweet potato and method for comprehensively utilizing component thereof |
| CN104804467A (en) * | 2015-05-15 | 2015-07-29 | 湖南农业大学 | Davidii foex pigment preparation method |
-
2015
- 2015-10-12 CN CN201510655494.1A patent/CN105199425A/en active Pending
Patent Citations (6)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| JPH01132344A (en) * | 1988-10-21 | 1989-05-24 | T Hasegawa Co Ltd | Food and drink |
| CN101186757B (en) * | 2007-12-24 | 2010-10-06 | 钱生球 | Method for producing purple sweet potato haematochrome |
| CN101735644A (en) * | 2009-11-20 | 2010-06-16 | 苏州大学 | Method for extracting ipomoes batatas L. anthocyanidins |
| CN101735647A (en) * | 2009-12-29 | 2010-06-16 | 威海华彩盛业生物科技有限公司 | Method for extracting ipomoes batatas L. pigment |
| CN102206426A (en) * | 2011-03-23 | 2011-10-05 | 武汉普赛特膜技术循环利用有限公司 | Comprehensive utilization of pigment of purple sweet potato in purple sweet potato and method for comprehensively utilizing component thereof |
| CN104804467A (en) * | 2015-05-15 | 2015-07-29 | 湖南农业大学 | Davidii foex pigment preparation method |
Non-Patent Citations (1)
| Title |
|---|
| 杨国才等: "柠檬酸-磷酸氢二钠缓冲液法提取紫甘薯花青素的工艺研究", 《南方农业学报》 * |
Cited By (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN105503981A (en) * | 2015-12-01 | 2016-04-20 | 大兴安岭林格贝寒带生物科技股份有限公司 | Method for extracting and separating anthocyanin from red cabbage |
| CN109588568A (en) * | 2018-12-18 | 2019-04-09 | 南京农业大学 | A method of procyanidine is added by daily ration and improves sow reproductive performance |
| CN109577031A (en) * | 2018-12-27 | 2019-04-05 | 河北科技大学 | Using vegetable colour to the method for textile dyeing |
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Application publication date: 20151230 |