Patent Application
20240058325
The invention generally relates to Plant Resources. More specifically, it pertains to an improved industrial process for the extraction of Alpha yohimbine from Rauwolfia species and the extract thereof. The process produces alpha yohimbine with a higher yield in a faster and more economical manner, as compared to prior art processes.
Rauwolfia is a genus of Evergreen trees and shrubs belonging to dogbane or Apocynaceae family. Rauwolfia genus comprises of about 100 species. Plants belonging to genus of Rauwolfia are native to tropical or sub-tropical regions of world including Europe, Africa, Asia, Australia, and the Central and South America. Rauwolfia typically grows at a temperature of about 10-40° C. Rauwolfia species are of great medicinal and economic importance because of the presence of different Phytochemicals including alcohols, sugars and glycosides, Flavonoids, Steroids, tannins and Alkaloids.
All parts of Rauwolfia plant including, roots, stems and leaves comprises of Indole Alkaloids. The major Indole Alkaloids present in Rauwolfia are ajmalidine, ajmaline, aricine, canescine, isoserine, serpentine. Serpentitnine, reserpiline, reserpine, Yohimbine, Yohimbinine etc.
https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4566472/).
In India, about 8 species of genus Rauwolfia are cultivated. Rauwolfia serpentina is also known as Indian Snakeroot or Sarpagandha. It consists of various alkaloids and is most commonly used in the field of medicine. It is used for the treatment of hypertension, psychotic disorders, intestinal disorders, etc. It is also used to treat Poisonous insect stings and snake bites. Rauwolfia canescens/Rauwolfia tetraphylla also known as devil-pepper, also contain numerous phytochemicals and is of great medicinal importance. Different species of Rauwolfia contain different amounts of phytochemicals.
Alpha Yohimbine—Yohimbine is one of the major Indole/carboline alkaloid, whose major source is the bark of Pausinystalia species preferably Pausinystalia yohimbe (Corynanthe yohimbe). A remarkable amount of yohimbine is also present in Rauwolfia species, Pseudochinchona species etc.
The spatial arrangement of functional group (—OH) and properties of alpha yohimbine are different from those of yohimbine. Alpha Yohimbine is one of the diastereoisomers of alkaloid yohimbine.
Various methods are present for the extraction of alpha yohimbine from Rauwolfia species, such as:
Most of prior art methods involve the use of sophisticated techniques or equipment such as Column Chromatography. Reverse phase chromatography, HPLC coupled with ion trap mass spectrometry etc. which are quite complex, time consuming and not economical on industrial scale.
Further the prior art methods are resulting in low yield of the product despite use of expensive solvents and a number of steps.
Also the previous process involves the use of a number of expensive solvents/reagents such as oxalic acid, tartaric acid, ethyl acetate etc.
Present invention overcomes these drawbacks and offers technical and economic advantages not offered by any of the prior art processes.
Process of present invention gives much higher yield than prior art processes, due to use of less number of solvents/reagents and process steps.
Use of less number of process steps help to reduce handling losses as suffered in previous processes, thus increasing yield.
Process of present invention is much more economical due to following reasons:—
This above invention discloses a method for the extraction of alpha yohimbine from the roots, stem and leaves of Rauwolfia species and determination of the purity of the compound using HPLC. The above invention also belongs to the same applicant as of present invention. It is the closest Prior art.
The process of this prior art patent uses a precipitation method including alternate steps of acidification and alkalization along with organic solvents. Aromatic non-polar solvent toluene was used for the process of extraction by maintaining 40-50° C. Temperature and 9 to 9.5 pH by addition of alkali followed by the extraction using an acid preferably Tartaric acid. Extraction is followed by alkalization and extraction with chloroform for the purification of the product. Methyl alcohol was used for the precipitation of the extracted product. Other organic acids such as tartaric acid, oxalic acid were used for lowering of the pH at various steps of acidification.
The present invention is by same applicant as with improvements, which have resulted in a new process, which is faster, more economical and results in much higher yield of about 50-65% more as compared to prior art process.
The present invention or process is using Alcoholic solvent “Methanol” instead of using Aromatic solvent toluene at normal pH range and 40-55° C. temperature which is basically enhancing the yield of the final extract (50-65%) increase in yield). The above invention involves multiple acidification and alkalization steps for extraction as well as purification of the final product which are complex and require more man hours, whereas in the present invention, the number of steps are less as after organic solvent alkali extraction, the present invention involves the extraction using an organic solvent that is Chloroform without the use of any acid. The process of acidification is not involved in the present invention, which reduces handling losses of chemicals, making process more economical than existing process. In already existing process, organic acids such as tartaric acid, oxalic acid, etc. were used for lowering pH whereas in the present process, these are not used at all. The present invention also eliminates the use of ethyl acetate during purification. Higher yield i.e. about 50-65% increase is obtained using the new process as compared to the already existing process, which has considerable industrial importance.
for Extracting Alpha Yohimbine (Rauwolscine)
of Alpha yohimbine from Rauwolfia species”-
From Rauwolfia Species”- Pawan Kumar Goel
The above invention discloses a process for obtaining antipsychotic canescens extracts and alkaloids including alpha yohimbine from the leaves of Rauwolfia tetraphylla. The above invention uses organic solvent alkali and acidic extraction steps for the extraction of the compound and uses column chromatography for the separation of the alpha yohimbine from the other compound or simply for the purification of the alpha yohimbine.
The present invention differs from the above invention, as in the above invention, separate technique was used for the extraction that is organic solvent acid and alkali extraction and separate technique was used for the purification of the compound that is column chromatography whereas in present invention, the compound is being purified using acid base extraction technique without the use of column chromatography. The present invention results in high yield of the compound as compared to the already existing invention. Also the present invention involves the extraction of alpha yohimbine only for the commercial use whereas the above invention involves the extraction of other alkaloids also along with alpha yohimbine.
The above invention discloses a method for the production of yohimbine hydrochloride from the yohimbe (Pausinystalia yohimbe) barks. The method involves the use of organic solvents and various acids for the extraction. Vacuum drying and resin column methods were used for the drying and concentration of the product.
The present invention involves the extraction of Alpha-yohimbine from a completely different species that is Rauwolfia tetraphylla hence, is completely different from the above invention and also the present invention does not involve the use of critical and time step methods such as resin column chromatography, hence is simple and economically feasible.
The above publication discloses a method for extraction and quantitative determination of yohimbine from different parts of Rauwolfia species. In above publication, the roots, stems and leaves were used and extraction was carried out using organic solvent methanol and by centrifugation and the solvent was further removed using filtration only. Quantitative determination was conducted using HPTLC and it was observed that 6.11% of yohimbine was present in leaves of Rauwolfia tetraphylla and in stems and roots, it was absent.
The present invention discloses a method for the extraction of alpha yohimbine using organic solvent acidic and basic extraction method and acid-base purification method from Rauwolfia species. The present invention produces high yield and high purity of alpha yohimbine. The present invention differs from the above invention as in above invention the extraction was carried out using organic solvent only whereas in the present invention, precipitation method along with use organic solvents, acid and base was used for extraction with alternate washing steps using halogenated solvent (Chloroform). Also the present invention is cost effective as it does not require the use of certain sophisticated equipments such as column.
The above publication discloses a simple isocratic HPLC method for quantification of α-yohimbine, isoreserpiline and tetrahydroalstonine. Samples were analysed by reverse phase chromatography on a Waters spherisorb ODS2 column using isocratic elution with acetonitrile containing 0.1% TEA and water containing 0.1% TFA (35:65, v/v) at a flow rate of 1 mL/min, a column temperature of 30 C and UV detection at 210 nm.
The present invention differs from the above publication as in the above invention the analyses of samples were done using certain complex and sophisticated methods and equipment such as reverse phase chromatography and Detection of extracted compounds was carried out using UV detection technique. Also, the above process is complicated as it uses a lot of solvents whereas the present invention uses only few solvents and hence is simple, convenient, commercially economic and less time-consuming process and is producing the product with high yield and purity.
The above study deals with the extraction and evaluation of Indole alkaloids from Rauwolfia serpentina. In the above process, Indole alkaloids were extracted using ethanol at 25° C. for about 12 hours. Extraction is followed by phytochemical screening, which was conducted using various tests, including Molisch's test, Fehling's test. Tannins test etc. And qualitative assessment was carried out using TLC and HPLC methods. Also anti-proliferative and antimicrobial activity of extract of Rauwolfia serpentine was detected. 11.27% crude extract was obtained in the above process.
The present invention discloses the extraction of alpha yohimbine from Rauwolfia species using organic solvent methanol and alternative acidification and alkalization steps along with precipitation of the extracted compound hence is completely different from the above invention. Also, the present invention is simple and not using any complex chromatography technique and chemicals, hence is cost effective and also producing quite high yield of alpha yohimbine as compared to the already existing processes.
The above publication discloses, a validated, rapid, sensitive and selective ultra HPLC coupled with hybrid triple quadrapole linear ion trap mass spectroscopy method in multiple reaction monitoring mode for simultaneous determination of bioactive MIAs in ethanolic extract of seven Rauwolfia species. The powdered plant materials in above process were extracted using ethanol by 30 minutes sonication using an ultra-sonic water bath. HPLC combined with hybrid triple quadrapole linear on trap mass spectroscopy technique was used for the analysis of the extracted alkaloids from seven Rauwolfia species.
The present invention differs from the already existing process because present invention does not use complex methodology and techniques such as sonication and chromatography for extraction and analysis of extract and also the extraction in present invention was conducted using organic solvent methanol along with use of acid base and other precipitation agents and halogenated solvents such as chloroform.
https://www.amazon.in/Team-Six-Supplements-Alpha-Yohimbine/dp/801LXDJ207
The above commercialized product was extracted from the bark of Rauwolfia vomitoria and is used as a dietary supplement, whereas the product disclosed in above invention as extracted from root, stem and leaves of Rauwolfia tetraphylla.
The main object of the present invention is to disclose an improved process for the extraction of alpha yohimbine from Rauwolfia species resulting in higher yield.
One more object of the present invention is to disclose simple and economically feasible industrial process for the extraction of alpha yohimbine from Rauwolfia species.
Still another object is to disclose that the extract obtained by the process of present invention contains yohimbine hydrochloride as a marker compound in analytically detectable amounts of more than 0.1% of the extract.
The present invention discloses an improved process for the extraction of alpha yohimbine from leaves of Rauwolfia species, preferably Rauwolfia canescens/Rauwolfia tetraphylla. The process of present invention gives, about 50-65% increased yield of alpha yohimbine i.e. 600-670 mg/100 g dry leaves as compared to prior art processes. Further, the extract obtained by the process of present invention contains yohimbine hydrochloride as a marker compound in analytically detectable amounts of more than 0.1% of the extract.
The extraction process is simple, convenient and commercially economical as it does not involve the use of sophisticated methods and techniques such as chromatography and mass spectrometry. Also, the disclosed process uses lesser number of solvents, which helps in the elimination of handling losses being suffered in the old processes. The disclosed process involves specific extraction steps such as organic solvent extraction, acidification, alkalization etc. The disclosed process involves the use of polar alcoholic solvent “Methanol” instead of non-polar aromatic solvent “Toluene” which results in production of extract with higher yield. Moreover, Carbon treatment is being used to remove color from the sample.
The following description is of best-contemplated mode of carrying out the invention. The description is made for the purpose of illustrating the general principles of the invention and should not be taken in a limiting sense.
The present invention discloses an improved commercially viable industrial process for the extraction of alpha yohimbine form the leaves of Rauwolfia canescens/Rauwolfia tetraphylla. The process disclosed in present invention produces remarkable higher yield of the final material (About 50-65%) increased yield) i.e. 600-670 mg/100 g of dry leaves (Yield in prior art process was 400 mg/100 g of dry leaves. Also, the improved process is commercially economical along with quite simple methodology.
Extraction is the first step to separate the desired natural constituents from the parts of plants. Various methods of natural product extraction are available including, organic solvent extraction, distillation, pressing, sublimation, etc. out of these methods, the simplest and most widely used technique for natural product extraction is “Solvent Extraction” technique. The basic principle associated for extraction with solvent extraction process is that, the solvent penetrates into the solid matrix which dissolves the desired solute depending upon the polarity of the solvent based on the law of similarity and known fact of chemistry that ‘Like dissolves like’ that is the polar solvents will dissolve the polar compounds and non-polar solvents will dissolve non polar compounds. Hence solvents are usually selected on the basis of compound of interest to be extracted.
For the extraction of polar compounds polar solvents are being used whereas non polar or slightly polar compounds are extracted with the help of non-polar solvents. Once, the solvent will dissolve the solute, the solute will diffuse out of the solid matrix, further the extracted solutes are collected and purified. The purification is usually carried out to obtain high purity compound. The extraction process can be coupled with High Performance Liquid chromatography in order to obtain highly pure product along with high yield of the desired product.
Other than polarity, particle size, pH and Temperature are the other factors that affect the overall process of extraction. Smaller or finer the particle size, better will be the extraction. Usually, high temperatures increase the solubility and diffusion of the solute, but if a specific range of temperature exceeds, it results in the loss of the solvent which as a result produces the extract with no. of undesirable impurities. pH that is acidic or basic nature of solvent can also affect the extraction process as the addition of acid or base to organic solvent can results in the separation of desired secondary metabolites.
The extraction procedure involves various steps. Some of the solvents can be changed without departing from the scope of the invention. Three different approaches can be used which are illustrated as Examples 1 and 2 below:
Process involves following steps:
indicates data missing or illegible when filed
It is clear from the above table that purity of product obtained by present process (Lot 01 and 02) is almost the same as that of purity of product prepared by earlier prior art process of the applicant himself. However, the present process is economical, faster and results in higher yield of 50-65% than earlier process. Additionally, the extract obtained by the process of present invention contains yohimbine hydrochloride as a marker compound in analytically detectable amounts of more than 0.1% of the extract.
The present invention discloses an improved process for the extraction of Alpha yohimbine from Rauwolfia species i.e. Rauwolfia canescens/Rauwolfia tetraphylla. The improved process results in production of compound with 50-65% more yield as compared to already existing processes. The average yield from already existing process was 400 mg/100 g dry leaves (4 Kg/Ton dry leaves) whereas the yield from proposed process has varied from 6.0 to 6.7 Kg/Ton of dry leaves of plant Rauwolfia species which was not obtained in any other processes. The Novelty lies in the use of methanol (polar molecule) instead of tolune (non-polar molecule) and also acid-base steps for the extraction of desired compound. Also, use of lesser number of steps and lesser number of solvents/reagents makes the improved process more economical than prior art processes.
The inventive step of the present invention lies in the disclosure of method which produces quite high yield of alpha yohimbine—about 50-65% increased yield as compared to other methods, used for the extraction process. Inventive step in the terms of technical advancement over prior art, lies in use of methanol (polar molecule) instead of toluene (non-polar), elimination of use of organic acids and acidification and alkalization steps to achieve high yield and purity of desired extract. Methanol being a highly polar solvent, extracts all the alkaloids and since a few steps have been omitted, the losses at these steps have been minimized, thus increasing the yield at final stage. The disclosed process is commercially economical as lesser number of organic solvents and reagents are being used.
Alpha yohimbine is of great medicinal importance. It is used to treat erectile dysfunction and promote weight loss. It is also used to boost mood, to improve athletics' performance, to treat Angina, diabetic neuropathy etc. Moreover, the process used in present invention for extraction of alpha yohimbine is simple and commercially economical. Hence the present invention is of great industrial importance.
| Number | Date | Country | Kind |
|---|---|---|---|
| 202111012485 | Mar 2021 | IN | national |
| Filing Document | Filing Date | Country | Kind |
|---|---|---|---|
| PCT/IN2021/050506 | 5/5/2021 | WO |
