# US 12,767,970 B2 · Methods and compositions for melanin synthesis > Agents: this data is available as JSON from the patentgrep API and MCP server — see https://patentgrep.com/llms.txt - Number: US 12,767,970 B2 - Document ID: US12767970B2 - Status: Granted - Kind: B2 - Application number: 18/209,263 - Filed: 2023-06-13 - Published: 2024-01-04 - Granted: 2026-10-20 - Earliest claimed: 2021-01-07 - Assignees: Ramot at Tel-Aviv University Ltd. - Applicants: Ramot At Tel Aviv University Ltd. - Inventors: Ayala Lampel; Tlalit Massarano; Avigail-Elah Baruch Leshem - Web page: https://patentgrep.com/patent/US12767970B2 ## CPC - C12P 21/02: having a known sequence of two or more amino acids, e.g. glutathione - C12P: FERMENTATION OR ENZYME-USING PROCESSES TO SYNTHESISE A DESIRED CHEMICAL COMPOUND OR COMPOSITION OR TO SEPARATE OPTICAL ISOMERS FROM A RACEMIC MIXTURE {(brewing of beer C12C; producing vinegar C12J; producing specific peptides or proteins C07K; producing enzymes C12N9/00; DNA or RNA concerning genetic engineering, vectors, e.g. plasmids, or their isolation, preparation or purification C12N15/00; measuring or testing processes involving enzymes or microorganisms C12Q; measuring or testing processes involving nucleic acid amplification reactions C12Q1/6844; fermentation processes to form a food composition, A21 or A23; compounds in general, see the relevant compound class, e.g. C01, C07)} ## Abstract The invention relates to a composition comprising a polymer—based droplet, wherein the polymer—based droplet comprises one or more of melanin, a tyrosine substrate with a cleavable protecting group on the side chain, tyrosine, tyrosinase and any combination thereof. The composition further relates to a method of synthesizing melanin comprising the steps of: obtaining a polymer—based droplet containing a tyrosine substrate with a cleavable protecting group on the side chain (protected tyrosine) or a polymer—based droplet containing tyrosine (unprotected); wherein the tyrosine is oxidized by tyrosinase and polymerizes so as to obtain melanin. ## Claims ### Claim 1 A composition comprising a polyethylene glycol (PEG)/dextran aqueous two-phase system (ATPS) comprising a PEG rich continuous phase and dextran-rich droplets dispersed in the PEG-rich continuous phase, wherein: the dextran-rich droplets comprise a tyrosinase and a tyrosine substrate optionally protected with a cleavable protecting group on a side chain; the ATPS is based on 5-10 w/w % PEG and 10-20 w/w % dextran; the continuous PEG-rich phase comprises a higher concentration of PEG than of dextran and the dextran-rich droplets comprise a higher concentration of dextran than of PEG; the tyrosine substrate and the tyrosinase preferentially partition into the dextran-rich droplets; and when the tyrosine is unprotected, the unprotected tyrosine is enzymatically oxidized by the tyrosinase to form melanin within the dextran-rich droplets. ### Claim 2 (depends on 1) The composition of claim 1, wherein the cleavable protecting group on a side chain of the tyrosine is photocleavable or chemically cleavable. ### Claim 3 (depends on 1) The composition of claim 1, wherein the protected tyrosine is ortho-nitrobenzyl tyrosine (ONB-Y). ### Claim 4 (depends on 1) The composition of claim 1, wherein the size of the droplet is between 100 nm to 200 μm. ### Claim 5 (depends on 1) The composition of claim 1, wherein the composition is a dermal composition or a cosmetic composition and comprises a dermally or cosmetically acceptable carrier. ### Claim 6 (depends on 1) The composition of claim 1, wherein the dextran has a molecular weight of about 10 kDa. ### Claim 7 (depends on 6) The composition of claim 6, wherein the PEG has a molecular weight of about 8 kDa. ### Claim 8 (depends on 1) The composition of claim 1, wherein the tyrosinase is enriched in the droplets by at least 5-fold compared to the continuous phase. ### Claim 9 (depends on 1) The composition of claim 1, wherein the melanin formed in the dextran-rich droplets exhibits reduced aggregation relative to melanin formed in a homogeneous aqueous phase. ### Claim 10 A method of synthesizing melanin comprising the steps of: preparing an aqueous two-phase system (ATPS) by dissolving 5-10 w/w % polyethylene glycol (PEG) and 10-20 w/w % dextran in an aqueous solution comprising a tyrosine substrate optionally protected with a cleavable protecting group on a side chain; centrifuging the ATPS to create an upper and a lower phase; collecting the upper phase comprising dextran-rich droplets dispersed in a PEG-rich continuous phase; adding tyrosinase to the collected upper phase; and waiting for the appearance of a dark brown color, indicating that the tyrosine substrate was enzymatically oxidized by the tyrosinase to form melanin within the dextran-rich droplets, wherein: if protected tyrosine is used, the method further comprises, following the addition of the tyrosinase, a step of cleaving the cleavable protecting group on the side chain of the protected tyrosine so as to obtain unprotected tyrosine; the PEG-rich continuous phase comprises a higher concentration of PEG than of dextran and the dextran-rich droplets comprise a higher concentration of dextran than of PEG; and the tyrosine substrate and the tyrosinase preferentially partition into the droplets. ### Claim 11 (depends on 10) The method of claim 10, wherein the protected tyrosine is ortho-nitrobenzyl tyrosine (ONB-Y). ### Claim 12 (depends on 10) The method of claim 10, wherein the cleaving is by irradiation. ### Claim 13 (depends on 10) The method of claim 10, wherein the irradiation is by UV irradiation.