# US 8,784,896 B2 · Antioxidant therapy with a whole, leech saliva extract

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- Number: US 8,784,896 B2
- Document ID: US8784896B2
- Status: Granted
- Kind: B2
- Application number: 13/624,860
- Filed: 2012-09-21
- Published: 2014-03-20
- Granted: 2014-07-22
- Earliest claimed: 2012-09-17
- Assignees: Biopep Solutions, Inc.
- Applicants: Biopep Solutions, Inc.
- Inventors: Abbas Mohammad Ghawi; Ahmed Merzouk; Abdualrahman Abdualkader; Mohammad Alaama
- Web page: https://patentgrep.com/patent/US8784896B2

## CPC

- A61K 35/62: Leeches; Worms, e.g. cestodes, tapeworms, nematodes, roundworms, earth worms, ascarids, filarias, hookworms, trichinella or taenia
- A01K 67/30: Rearing or breeding invertebrates
- A61K 9/0014: {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)}
- A61K 31/282: Platinum compounds
- A61K 31/4745: condensed with ring systems having nitrogen as a ring hetero atom, e.g. phenantrolines (yohimbine derivatives, vinblastine A61K31/475; ergoline derivatives A61K31/48)
- A61K 31/555: containing heavy metals, e.g. hemin, hematin, melarsoprol
- A61K 38/21: Interferons {[IFN]}
- A61K 45/06: Mixtures of active ingredients without chemical characterisation, e.g. antiphlogistics and cardiaca
- A61P 3/10: for hyperglycaemia, e.g. antidiabetics
- A61P 31/04: Antibacterial agents
- A61P 31/12: Antivirals
- A61P 33/00: Antiparasitic agents
- A61P 35/00: Antineoplastic agents
- A61P 35/02: specific for leukemia
- A61P 39/06: Free radical scavengers or antioxidants
- Y02A 50/30: Against vector-borne diseases, e.g. mosquito-borne, fly-borne, tick-borne or waterborne diseases whose impact is exacerbated by climate change
- A61K: PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES (devices or methods specially adapted for bringing pharmaceutical products into particular physical or administering forms A61J3/00; chemical aspects of, or use of materials for deodorisation of air, for disinfection or sterilisation, or for bandages, dressings, absorbent pads or surgical articles A61L; soap compositions C11D)
- A01K: ANIMAL HUSBANDRY; AVICULTURE; APICULTURE; PISCICULTURE; FISHING; REARING OR BREEDING ANIMALS, NOT OTHERWISE PROVIDED FOR; NEW BREEDS OF ANIMALS
- A61P: SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
- Y02A: TECHNOLOGIES FOR ADAPTATION TO CLIMATE CHANGE

## Abstract

Methods are provided for isolating and using a whole-saliva leech extract. The methods can include feeding a phagostimulatory agent to a leech; inducing a regurgitation in the leech, the inducing including placing the leech in an environment having a temperature of less than about 0° C.; and, collecting an unrefined, whole saliva in the regurgitation of the cooled leech. The methods can include revitalizing the leech by warming it at a temperature ranging from about 5° C. to about 40° C. Stable, lyophilized, whole-saliva extracts of a leech are also provided, the extract having a stable activity when stored for use at a temperature below about −20° C., the extract maintaining at least 70% of the activity for at least 6 months. The extracts can be used to treat solid tumors, treat liquid tumors, treat diabetes, treat a viral disease, treat a parasitic disease, treat an antibacterial disease, or serve as an anti-oxidant.

## Claims

### Claim 1

A method of scavenging free radicals, comprising
contacting a free radical in a fluid with a whole, leech saliva extract; wherein,
the contacting includes scavenging the free radicals in the fluid; and,
the extract has a peptide molecular weight distribution with molecular weights of 3496 Daltons or greater wherein the extract is produced from a process comprising: feeding a phagostimulatory agent to a leech; inducing a regurgitation in the leech, the inducing including placing the leech in an environment having a temperature of less than 0° C. or about 0° C.; and, collecting an unrefined, whole saliva in the regurgitation of the cooled leech.

### Claim 2 (depends on 1)

The method of claim 1, wherein the collecting includes squeezing the leech to increase the amount of unrefined, whole saliva collected.

### Claim 3 (depends on 1)

The method of claim 1, further comprising revitalizing the leech by warming the leech in a water bath having a temperature ranging from about 5° C. to about 40° C.

### Claim 4 (depends on 1)

The method of claim 1, wherein the method further comprises creating a refined, whole-saliva extract; the creating including removing solid components from the unrefined, whole saliva.

### Claim 5 (depends on 1)

The method of claim 1, wherein the method further comprises lyophilizing separate volumes of the refined, whole saliva extract, the volumes not exceeding about 2 ml each.

### Claim 6 (depends on 1)

The method of claim 1, the leech being Hirudinaria manillensis.

### Claim 7

A method of scavenging free radicals, comprising contacting a free radical in a fluid with a whole, leech saliva extract, wherein the contacting includes scavenging the free radicals in the fluid; wherein the extract has a peptide molecular weight distribution with molecular weights of 3496 Daltons or greater, and wherein the extract is a lyophilized, whole saliva extract of a leech produced from a method comprising:
feeding a phagostimulatory agent to a leech;
inducing regurgitation in the leech, the inducing including placing the leech in an environment having a temperature ranging from about −5° C. to about 15° C.
collecting an unrefined, whole saliva in the regurgitation of the cooled leech;
removing solid components from the unrefined, whole saliva to create a refined, whole saliva; and,
lyophilizing separate volumes of the refined, whole saliva extract, the volumes not exceeding about 2 ml each.

### Claim 8 (depends on 7)

The method of claim 7, wherein the collecting includes squeezing the leech to increase the amount of unrefined, whole saliva collected.

### Claim 9 (depends on 7)

The method of claim 7, further comprising revitalizing the leech by warming the leech in a water bath having a temperature ranging from about 5° C. to about 40° C.

### Claim 10 (depends on 7)

The method of claim 7, the leech being Hirudinaria manillensis.

### Claim 11 (depends on 7)

The method of claim 7, wherein the collecting of the whole saliva yields a peptide molecular weight distribution that ranges from 4276 Daltons to 44386 Daltons.

### Claim 12 (depends on 7)

The method of claim 7, wherein the collecting of the whole saliva yields a peptide molecular weight distribution that ranges from 6289 Daltons to 14244 Daltons.

### Claim 13 (depends on 7)

The method of claim 7, wherein the collecting of the whole saliva yields a peptide molecular weight distribution that ranges from 10812 Daltons to 88210 Daltons.

### Claim 14 (depends on 7)

The method of claim 7, wherein the collecting of the whole saliva yields a peptide molecular weight distribution that ranges from 3496 Daltons to 88210 Daltons.

### Claim 15 (depends on 1)

The method of claim 1, wherein the fluid is in a subject in need of an antioxidant therapy, and the antioxidant therapy is used to treat an arthritis.

### Claim 16 (depends on 1)

The method of claim 1, wherein the fluid is in a subject in need of an antioxidant therapy, and the antioxidant therapy is used to treat an inflammatory disorder.

### Claim 17 (depends on 1)

The method of claim 1, wherein the fluid is in a subject in need of an antioxidant therapy, and the antioxidant therapy is used to treat a cardiovascular disease.

### Claim 18 (depends on 1)

The method of claim 1, wherein the fluid is in a subject in need of an antioxidant therapy, and the antioxidant therapy is used to treat atherosclerosis.

### Claim 19 (depends on 1)

The method of claim 1, wherein the extract is administered in combination with an absorption blocker in the treatment of cardiovascular disease.

### Claim 20 (depends on 1)

The method of claim 1, wherein the extract is administered in combination with a synthesis inhibitor in the treatment of cardiovascular disease.

### Claim 21 (depends on 1)

The method of claim 1, wherein the extract is administered in combination with a niacin-based drug in the treatment of cardiovascular disease.

### Claim 22 (depends on 1)

The method of claim 1, wherein the fluid is in a subject in need of an antioxidant therapy, and the antioxidant therapy is used to treat aging.

### Claim 23 (depends on 1)

The method of claim 1, wherein the fluid is in a subject in need of an antioxidant therapy, and the antioxidant therapy is used as a neuroprotective treatment.