the disclosed embodiments are directed to a rapid , inexpensive and easy - to - use therapeutic and regenerative agent that promotes healing , reduces inflammation and treats various afflictions in animals , among other things . the disclosed embodiments improve over the prior art by providing a simple , inexpensive and quick method for producing said therapeutic agent for mass production and transport to individuals and consumers . the disclosed embodiments also improve over the prior art by providing an agent that can be used as cell - culture to promote the growth of desired cells . the following detailed description refers to the accompanying drawings . whenever possible , the same reference numbers are used in the drawings and the following description to refer to the same or similar elements . while disclosed embodiments may be described , modifications , adaptations , and other implementations are possible . for example , substitutions , additions or modifications may be made to the elements illustrated in the drawings , and the methods described herein may be modified by substituting reordering , or adding additional stages or components to the disclosed methods and devices . accordingly , the following detailed description does not limit the disclosed embodiments . instead , the proper scope of the disclosed embodiments is defined by the appended claims . the claimed process produces an a - cellular product , derived from umbilical cord tissue and wharton &# 39 ; s jelly that contains cytokines , growth factors , peptides , signaling molecules , proteins , exosomes and anti - inflammatory and regenerative molecules that can be used by itself or combined with a variety of different products including but not limited to ; cream - base containing lipophilic agents and can be used to treat anti - inflammatory conditions in mammals ( both human and veterinary applications ), gel and agarose based products for the delivery of these factors to an external wound in mammals ( both human and veterinary applications ), can be aerosolized and used via a nebulizer to treat conditions of the lung , can be injected or administered systemically to treat a variety of conditions such as cardiac , neurologic , musculoskeletal , hepatic , n can be lyophilized or freeze - dried and reconstituted for long - term storage and use . the product can also be used as cell - culture to promote the growth of desired cells . the claimed process discloses methods of generating a therapeutic and regenerative product from umbilical cord tissue and wharton &# 39 ; s jelly tissue . in one embodiment , the claimed process provides means of creating a therapeutic and regenerative product useful for the treatment of inflammatory conditions , wounds and degenerative conditions by producing an extract of umbilical cord tissue and wharton &# 39 ; s jelly . many types of methods of creating a tissue extract may be used and chosen . in one embodiment , a method is selected from a group comprising ; 1 ) sonication ; 2 ) shearing by liquid flow ; 3 ) exploding by pressure ; 4 ) collision forces by impact of beads or paddles ; 5 ) cryogenic grinding ; 6 ) mortar and pestle ; 7 ) glass homogenizer ; 8 ) blender ; 9 ) rotor - stator ; 10 ) potter - elvehjem with ptfe pestle ; 11 ) french press ; 12 ) amalgamators for tubes ; 13 ) high throughput homogenizers ; 14 ) combination of the above methods , or any similar methods known in the art . in some embodiments , the tissue extract is administered directly into the patient ( human or animal ). it is well known in the art that preparation of the extract before administration may be performed by various means , for example , said extract may be filter sterilized or in some conditions concentrated or diluted . in one embodiment , the product can be directly administered by injection . in other embodiments , the tissue extract is used as an active ingredient for the generation of a pharmaceutical formulation . this may comprise administration of the tissue extract therapeutic agent alone , or by way of known pharmaceutical formulations , including tablets , capsules , or elixirs for oral administration , suppositories for rectal administration , sterile solutions or suspensions for parenteral or intramuscular administration , liposomal or encapsulated formulations , formulations wherein the therapeutic agent is alone or conjugated to a delivery agent or vehicle , and the like . it is possible that therapeutic entities of the product will be administered with suitable carriers , excipients , and / or other agents that are incorporated into formulations to provide improved transfer , delivery , tolerance , and the like . a multitude of appropriate formulations can be found in the formulary known to all pharmaceutical chemists : remington &# 39 ; s pharmaceutical sciences ( 15 . sup . th ed , mack publishing company , easton , pa . ( 1975 )), particularly chapter 87 by blaug , seymour , therein . these formulations include , for example , powders , pastes , ointments , jellies , waxes , oils , lipids , lipid ( cationic or anionic ) containing vesicles ( such as lipofectin . tm . ), dna conjugates , anhydrous absorption pastes , oil - in - water and water - in - oil emulsions , emulsions carbowax ( polyethylene glycols of various molecular weights ), semi - solid gels , and semi - solid mixtures containing carbowax . any of the foregoing mixtures may be appropriate in treatments and therapies in accordance with the claimed subject matter , provided that the active ingredient in the formulation is not inactivated by the formulation and the formulation is physiologically compatible and tolerable with the route of administration . see also powell et al . “ compendium of excipients for parenteral formulations ” pda j pharm sci technol 52 : 238 - 311 ( 1998 ). in one embodiment , one or more agents of the claimed process are nanoencapsulated into nanoparticles for delivery . the nanoencapsulation material may be biodegradable or non - degradable . the nanoencapsulation materials may be made of synthetic polymers , natural polymers , oligomers , or monomers . synthetic polymers , oligomers , and monomers include those derived from polyalkyleneoxide precursor molecules , such as poly ( ethylene oxide ) ( peo ), poly ( ethylene glycol ) ( peg ) and copolymers with poly ( propylene oxide ) ( peg - co - ppo ), poly ( vinyl alcohol ) ( pva ), poly ( vinylpyrrolidone ) ( pvp ), poly ( ethyloxazoline ) ( peox ), polyaminoacids , and pseudopolyamino acids , and copolymers of these polymers . copolymers may also be formed with other water - soluble polymers or water insoluble polymers , provided that the conjugate is water soluble . an example of a water - soluble conjugate is a block copolymer of polyethylene glycol and polypropylene oxide , commercially available as a pluronic surfactant ( basf ). natural polymers , oligomers and monomers include proteins , such as fibrinogen , fibrin , gelatin , collagen , elastin , zein , and albumin , whether produced from natural or recombinant sources , and polysaccharides , such as agarose , alginate , hyaluronic acid , chondroitin sulfate , dextran , dextran sulfate , heparin , heparin sulfate , heparan sulfate , chitosan , gellan gum , xanthan gum , guar gum , water soluble cellulose derivatives , and carrageen . these polymers are merely exemplary of the types of nanoencapsulation materials that can be utilized and are not intended to represent all the nanoencapsulation materials within which entrapment is possible . in one embodiment , the therapeutic agent is administered in a topical formulation . topical formulations are useful in the treatment of conditions associated with dermal diseases and joint disorders / joint pain . for example , topical administration of the tissue extract may contain aqueous and non - aqueous gels , creams , multiple emulsions , microemulsions , liposomes , ointments , aqueous and non - aqueous solutions , lotions , aerosols , skin patches , hydrocarbon bases and powders , and can contain excipients such as solubilizers , permeation enhancers ( e . g ., fatty acids , fatty acid esters , fatty alcohols and amino acids ), and hydrophilic polymers ( e . g ., polycarbophil and polyvinylpyrolidone ). in one embodiment , the pharmaceutically acceptable carrier is a liposome or a transdermal enhancer . topical formulations of the product may include a dermatologically acceptable carrier , e . g ., a substance that is capable of delivering the other components of the formulation to the skin with acceptable application or absorption of those components by the skin . the carrier will typically include a solvent to dissolve or disperse the therapeutic agent , and , optionally one or more excipients or other vehicle ingredients . carriers useful in accordance with the topical formulations of the product may include , by way of non - limiting example , water , acetone , ethanol , ethylene glycol , propylene glycol , butane - 1 , 3 - diol , acrylates copolymers , isopropyl myristate , isopropyl palmitate , mineral oil , butter ( s ), aloe , talc , botanical oils , botanical juices , botanical extracts , botanical powders , other botanical derivatives , lanolin , urea , petroleum preparations , tar preparations , plant or animal fats , plant or animal oils , soaps , triglycerides , and keratin ( s ). topical formulations of the product are prepared by mixing a compound with a topical carrier . in other embodiments , moisturizers or humectants , sunscreens , fragrances , dyes , and / or thickening agents such as paraffin , jojoba , paba , and waxes , surfactants , occlusives , hygroscopic agents , emulsifiers , emollients , lipid - free cleansers , antioxidants and lipophilic agents , may be added to the topical formulations . a topical formulation may be designed to be left on the skin and not washed shortly after application . alternatively , the topical formulation may be designed to be rinsed off within a given amount of time after application . in one aspect , potency of the tissue extract product may be quantified by assessing protein concentration . for quantification of anti - inflammatory and regenerative activity , the term “ inflammation ” will be understood to include any condition characterized by a localized or a systemic protective response , which may be elicited by physical trauma , infection , chronic diseases , such as those mentioned above , and / or chemical and / or physiological reactions to external stimuli ( e . g ., as part of an allergic response ). any such response , may be manifested by heat , swelling , pain , redness , dilation of blood vessels and / or increased blood flow , invasion of the affected area by white blood cells , loss of function and / or any other symptoms known to be associated with inflammatory conditions . the term “ inflammation ” will thus also be understood to include any inflammatory disease , disorder or , any condition that has an inflammatory component associated with it , and / or any condition characterized by inflammation as a symptom , including , inter alia , acute , chronic , ulcerative , specific , allergic and necrotic inflammation , and other forms of inflammation . the term thus also includes inflammatory pain . for the practice of the claimed subject matter supernatants generated by umc & amp ; wharton &# 39 ; s jelly tissue extracts may be administered to the patient in an injection solution , which may be saline , lactated ringers solution , mixtures of autologous plasma together with saline , or various concentrations of albumin with saline . ideally ph of the injection solution is from about 6 . 4 to about 8 . 3 , optimally 7 . 4 . excipients may be used to bring the solution to isotonicity such as , 4 . 5 % mannitol or 0 . 9 % sodium chloride , ph buffers , such as sodium phosphate . other pharmaceutically acceptable agents can also be used to bring the solution to isotonicity , including , but not limited to , dextrose , boric acid , sodium tartrate , propylene glycol , polyols ( such as mannitol and sorbitol ) or other inorganic or organic solutes . injection can be performed systemically , or more specifically , via routes of administration selected from ; a ) orally ; b ) intravenously ; c ) intramuscularly ; d ) intraperitoneally ; e ) intrathecally ; f ) alimentarily ; g ) intraspinally ; h ) intra - articularly ; i ) intra - joint ; j ) subcutaneously ; k ) buccally ; l ) vaginally ; m ) rectally ; n ) dermally ; o ) transdermally ; p ) ophthalmically ; q ) auricularly ; r ) mucosally ; s ) nasally ; t ) tracheally ; u ) bronchially ; v ) sublingually ; w ) intranodally ; x ) by any parenteral route ; and y ) via inhalation . the manufacturing of the disclose product may entail the collection of umbilical cord tissue . the umbilical cord is collected after the new offspring is delivered by clamping the cord proximal to the offspring and then by cutting the cord . the freshly cut umbilical cord is then placed into a container containing buffered saline solution or similar buffered salt solution for transportation to the processing laboratory . the process may proceed by incubating the umbilical cord in a solution containing antibiotics for sterilization at a temperature not exceeding 50 or 70 degrees centigrade . the umbilical cord is washed in salt buffer about three times and the vasculature can be or may not be removed by micro - dissection and discarded . the dissected tissue is then washed about three more times in a salt buffer to remove any residual blood and minced into small fragments ( less than 1 mm in size ) using a blender ( or any other method known in the art ), resulting in an aqueous solution . the aqueous solution , including minced tissue , is then re - suspended in a salt buffer and subjected to sonication . this step uses blasts of ultrasonic sound waves to disrupt the cells and tissue to facilitate the release of the different cytokines , peptides and peptide growth factors that will be the active components in the treatment of inflammation . after sonication the aqueous solution ( including disrupted tissue and buffer ) is transferred into centrifuge tubes and centrifuged at 1 , 000 - 10 , 000 rpm for 10 minutes to pellet all the tissue and cellular debris and to separate the aqueous solution into a soluble component and a non - soluble component . the supernatant containing all the cytokines , peptides , signaling molecules , and peptide growth factors is transferred into a new centrifuge tube and centrifuged again at 1 , 000 - 10 , 000 rpm for 10 minutes to complete the removal and sedimentation of the tissue and cellular debris . the supernatant is then passed through a 100 μm cell strainer to remove any left - over cellular debris , then it is passed through a 0 . 22 μm polypropylene syringe filter to sterile filter the supernatant . i . e ., the process filtrates and discards the non - soluble component of the aqueous solution from the centrifuge through a 0 . 2 micron filter . next , the protein concentration of the soluble component is measured to insure at least about 0 . 05 microgram of total protein extract . the process may further comprise mixing the soluble component with liposomes , resulting in a mix , mixing the mix with cream as a vehicle for application to a mammal , so as to produce a mixed cream , and depositing the mixed cream in a container for transport to users . in a first example of experimental results , a three - year - old female horse suffered a severe burn during the removal of a wart on the back of the right front foot . the wound persisted for 2 months without any substantial healing . the pictures shown in fig2 were taken the first week after the burn occurred . treatment with the uct wharton &# 39 ; s jelly extract ( i . e ., the product produced by the claimed subject matter ) began on may 9 , 2016 with the extract being applied directly to the wound 3 times a week . the wound was wrapped and measurements were taken . fig3 and 5 show photographs of the wound at various dates indicated , as well as the size of the wound , which clearly shows that the wound heals over time , due to the use of the product produced by the claimed subject matter . the leftmost photograph of fig3 , for example , shows a 5 - 9 - 2016 photograph of a wound sized 12 cm × 4 . 5 cm × 1 . 25 cm , while the rightmost photograph of fig3 shows a 5 - 22 - 2016 photograph of a wound sized 7 cm × 3 . 75 cm × 0 . 5 cm . in a second example of experimental results , a horse that suffered a severe laceration to its hoof area . the product produced by the claimed subject matter was combined in a moisturizer like cream and applied to the wound 2 × daily . fig6 shows photographs of the wound at various dates indicated , as well as the size of the wound , which clearly shows that the wound heals over time . the leftmost photograph of fig6 , for example , shows a 9 am jan . 9 , 2017 photograph taken in mert , tex . of a wound before treatment with the product produced by the claimed subject matter , while the bottom photograph of fig6 shows a 4 pm jan . 20 , 2017 photograph taken in mert , tex . of said wound after treatment with the product produced by the claimed subject matter , clearly showing improvement . in a third example of experimental results , a 7 - year - old male horse that broke his left fetlock at age 2 . he has suffered from chronic suspensory branch lesions with scar tissue formation . the horse has been bandaged constantly and has had corrective shoeing performed to improve soundness with no improvements . the product produced by the claimed subject matter was applied to this horse 2 × a day for 7 days with the following results demonstrating the decrease in inflammation in a very short period . fig7 shows the decrease in inflammation due to the wound exhibited by the horse over time . fig7 shows that the wound was about 32 units in diameter on day 1 , but shrank to about 26 units in diameter at day 3 of the treatment . in a fourth example of experimental results , a six - year - old male horse that suffered an injury where the splint bones popped . the injury did not heal resulting in physical calcification of the splint bone to the cannon bone causing inflammation and sensitivity to the suspensory ligament . the product produced by the claimed subject matter was applied 2 × a day for 7 days . the injury occurred 4 months ago and was initially treated with surpass and poltice clay with no effect . this treatment was performed 4 months before the application of the extract . fig8 shows the decrease in inflammation due to the wound exhibited by the horse over time . fig8 shows that the wound was about 7 . 5 units in length on day 1 , but shrank to about 5 . 5 units in length at day 7 of the treatment . in a fifth example of experimental results , horse suffered a laceration above its eye that required sutures . the product produced by the claimed subject matter was administered after the sutures were performed 2 times a day . the results of wound closure were from 3 days application of the claimed product . fig9 shows photographs of the wound at different dates , which clearly shows that the wound heals over time . in a sixth example of experimental results , an 8 - month - old puppy exhibited an ulcer on its pectoral area that turned out to be a staph aureus infection ( confirmed by independent testing lab and an independent physician ). after four days of treatment with the product produced by the claimed subject matter and a wound cream , there was total clearance of the staph infection . fig1 shows photographs of the wound at different dates , which clearly shows that the wound heals over time . anti - microbial activity could be attributed to 2 possible mechanisms : the first is that the umbilical extract contains antimicrobial peptides that play a direct role in the antimicrobial activity , or secondly , there are numerous cytokines and immune signaling molecules present in the claimed produce that there is a massive influx of immune cells into the location were the cream has been administered . one possible mechanism of action of the product produced by the claimed subject matter is that it recruits endogenous stem cells to the location that it is applied to or injected into . see fig1 for a description of said mechanism , which includes the paracrine factors , and shows how the immunomodulatory effects , angiogenesis and recruitment of endogenous stem cells are processed for proliferation and differentiation . in an attempt to demonstrate this experimentally , msc migration assays were performed . mscs were seeded into culture containing a filter paper soaked in product produced by the claimed subject matter and a control was performed with the filter paper soaked in standard rpmi culture media . the cells in the product produced by the claimed subject matter media can be clearly seen migrating and filling in the area where the filter paper was at the 24 hour time point ( paper removed to show the cells ) and with the control media , rpmi , no cell migration can be seen . see fig1 and 13 . additionally it was observed that mscs proliferation was always greater in media supplemented with product produced by the claimed subject matter versus media supplemented with fbs . thus , the product produced by the claimed subject matter can be used as cell culture . although the subject matter has been described in language specific to structural features and / or methodological acts , it is to be understood that the subject matter defined in the appended claims is not necessarily limited to the specific features or acts described above . rather , the specific features and acts described above are disclosed as example forms of implementing the claims .
a method of preparation of a solution used for therapeutic and regenerative applications having a concentration of at least about . 05 microgram of total protein extract is disclosed . the method comprises the steps of : collecting an umbilical cord ; incubating the umbilical cord in a solution containing antibiotics for sterilization ; washing the umbilical cord in a buffered salt solution ; homogenizing the umbilical cord in the buffered salt solution so as to produce an aqueous solution including the buffered salt solution ; applying a sonicator at ultrasonic frequencies to the aqueous solution to further disrupt the aqueous solution ; placing the aqueous solution into a centrifuge to separate the aqueous solution into a soluble component and a non - soluble component ; filtrating and discarding the non - soluble component of the aqueous solution from the centrifuge through a 0 . 2 micron filter ; and measuring protein concentration of the soluble component to insure at least about 0 . 05 microgram of total protein extract .