Method for the microbial production of specific natural capsaicinoids

Patent metadata

Field	Value
Publication number	US-11946083-B2
Application number	US-202217821810-A
Country	US
Kind code	B2
Filing date	Aug 24, 2022
Priority date	Jul 19, 2016
Publication date	Apr 2, 2024
Grant date	Apr 2, 2024

Abstract

The present invention relates to the production of capsaicinoid compounds including Capsaicin and Nonivamide via microbial fermentation.

First claim

What is claimed is:

1. A biosynthetic method of making a glucoside of a capsaicinoid of interest comprising:
a) expressing an acyl-CoA synthetase (ACS) and capsaicin synthase (CS) in a transformed cellular system;
b) expressing a UDP-glucosyltransferase in the transformed cellular system, wherein the UDP-glucosyltransferase is a CaUGT2 from C. roseus and either is encoded by a DNA sequence at least 75% identical to SEQ ID NO: 5 and/or comprises an amino acid sequence at least 90% identical to SEQ ID NO: 6;
c) growing the cellular system in a medium; and,
d) producing the glucoside of a capsaicinoid of interest.

2. The biosynthetic method of claim 1 , wherein both the ACS and CS are cloned from a plant of the Capsicum genus.

3. The biosynthetic method of claim 2 , wherein the Capsicum genus plant is the ghost chili.

4. The biosynthetic method of claim 1 , wherein the ACS expressed is derived from a gene cloned from Arabidopsis based on LCAS4 or LCAS5.

5. The biosynthetic method of claim 1 , further comprising feeding a source material to said cellular system in addition to culture media.

6. The biosynthetic method of claim 5 , wherein said source material is selected from the group consisting of nonanoic acid, oil of pelargonium, octanoic acid, and decanoic acid.

7. The biosynthetic method of claim 5 , wherein said source material is selected from the group consisting of C6 to C12 hydrocarbons that are medium to long chain fatty acids.

8. The biosynthetic method of claim 5 , wherein the transformed cellular system further comprises an aminotransferase which is capable of catalyzing the conversion of vanillin to vanillylamine, and the source material further comprises at least one of vanillin and vanillyamine.

9. The biosynthetic method of claim 1 , wherein said transformed cellular system is selected from the group consisting of yeast, non-capsaicinoid producing plants, algae and bacteria.

10. The biosynthetic method of claim 9 , wherein said transformed cellular system is E.Coli.

11. The biosynthetic method of claim 1 , wherein the capsaicinoid of interest is selected from the group consisting of capsaicin, nonivamide, N-vanillylnonanamides, N-vanillylsulfonamides, N-vanillylureas, N-vanillylcarbamates, N[(substituted phenyl)methyl]alkylamides, methylene substituted N[(substituted phenyl)methyl]alkanamides, N[(substituted phenyl)methyll-cis-monosaturated alkenamides, N[(substituted phenyl)methyl]diunsaturated amides, 3-hydroxyacetanilide, hydroxyphenylacetamides, pseudocapsaicin, dihydrocapsaicin, nordihydrocapsaicin, homocapsaicin, homodihydrocapsaicin I, anandamide, piperine, zingerone, warburganal, polygodial, aframodial, cinnamodial, cinnamosmolide, cinnamolide, civamde, nonivamide, olvanil, N-oleylhomovanillamidia, isovelleral, scalaradial, ancistrodial, and any combinations or mixtures thereof.

12. The biosynthetic method of claim 1 , wherein the capsaicinoid of interest is capsaicin or nonivamide.

13. The biosynthetic method of claim 1 , wherein the ACS is encoded by a DNA sequence at least 75% identical to SEQ ID NO: 3.

14. The biosynthetic method of claim 1 , wherein the ACS comprises an amino acid sequence at least 90% identical to SEQ ID NO: 1.

15. The biosynthetic method of claim 1 , wherein the CS is encoded by a DNA sequence at least 75% identical to SEQ ID NO: 4.

16. The biosynthetic method of claim 1 , wherein the CS comprises an amino acid sequence at least 90% identical to SEQ ID NO: 2.

17. The biosynthetic method of claim 1 , wherein the ACS is encoded by a DNA sequence comprising the nucleotide sequence of SEQ ID NO: 3.

18. The biosynthetic method of claim 1 , wherein the ACS comprises the amino acid sequence of sequence of SEQ ID NO: 1.

19. The biosynthetic method of claim 1 , wherein the CS is encoded by a DNA comprising the nucleotide sequence of SEQ ID NO: 4.

20. The biosynthetic method of claim 1 , wherein the CS comprises the amino acid sequence of SEQ ID NO: 2.

Assignees

• Conagen Inc

Inventors

• Chen Hui
• Yu Xiaodan
• Zhou Lanlan
• Wang Hongxue
• Wang Min

Classifications

• C12P13/02 Primary

  Amides, e.g. chloramphenicol {or polyamides; Imides or polyimides; Urethanes, i.e. compounds comprising N-C=O structural element or polyurethanes (peptides C12P21/00 or C07K)} · CPC title

• A23L27/10

  Natural spices, flavouring agents or condiments; Extracts thereof · CPC title

• A61K9/0014

  Skin, i.e. galenical aspects of topical compositions (non-active ingredients are additionally classified in A61K47/00; A61K9/0009, A61K9/0021, A61K9/7015, A61K9/7023 take precedence; cosmetic preparations A61K8/00, A61Q; preparations for wound dressings or bandages A61L26/00) · CPC title

• C07C233/18

  having the carbon atom of the carboxamide group bound to a hydrogen atom or to a carbon atom of an acyclic saturated carbon skeleton · CPC title

• C07C233/20

  having the carbon atom of the carboxamide group bound to a carbon atom of an acyclic unsaturated carbon skeleton · CPC title