( 12 ) United States Patent
`Lundberg et al .
`
`US 10,842,813 B2
`( 10 ) Patent No .:
`( 45 ) Date of Patent :
`Nov. 24 , 2020
`
`US010842813B2
`
`( 54 ) COMPOSITIONS OF NITRATES AND
`METHODS OF USE THEREOF
`( 71 ) Applicant : Heartbeet Ltd. , Ashbocking ( GB )
`( 72 ) Inventors : Jon Lundberg , Djursholm ( SE ) ; Eddie
`Weitzberg , Stockholm ( SE )
`( 73 ) Assignee : HEARTBEET LTD , Asbocking ( GB )
`Subject to any disclaimer , the term of this
`( * ) Notice :
`patent is extended or adjusted under 35
`U.S.C. 154 ( b ) by 157 days .
`( 21 ) Appl . No .: 15 / 966,508
`Apr. 30 , 2018
`( 22 ) Filed :
`( 65 )
`Prior Publication Data
`May 2 , 2019
`US 2019/0125784 A1
`Related U.S. Application Data
`( 63 ) Continuation - in - part of application No. 12 / 528,794 ,
`filed as application No. PCT / SE2008 / 050212 on Feb.
`( Continued )
`Foreign Application Priority Data
`
`( SE )
`( SE )
`
`0700520
`0700729
`
`( 30 )
`Feb. 26 , 2007
`Mar. 22 , 2007
`( 51 ) Int . Cl .
`A61K 33/00
`A61K 9/00
`
`( 2006.01 )
`( 2006.01 )
`( Continued )
`( 52 ) U.S. CI .
`A61K 33/00 ( 2013.01 ) ; A23L 2/02
`CPC
`( 2013.01 ) ; A23L 2/52 ( 2013.01 ) ; A23L 33/105
`( 2016.08 ) ;
`( Continued )
`( 58 ) Field of Classification Search
`None
`See application file for complete search history .
`beetroot juice
`3-4 dl daily
`
`( 56 )
`
`EP
`EP
`
`4,702,922 A
`4,868,179 A
`
`References Cited
`U.S. PATENT DOCUMENTS
`10/1987 Wiesenberger
`9/1989 Cohn
`( Continued )
`FOREIGN PATENT DOCUMENTS
`
`11/1992
`0511587 A1
`2008661 A2 12/2008
`( Continued )
`
`OTHER PUBLICATIONS
`“ 4.2 Acute blood pressure ( BP ) lowering and
`AJ Webb et al . ,
`vasoprotective effects of beetroot juice : relationship to nitrite derived
`from dietary nitrate ” , Journal of Human Hypertension , vol . 21 , No.
`10 , pp . 834-835 . ( Year : 2007 ) . *
`( Continued )
`
`Primary Examiner - Bethany P Barham
`Assistant Examiner - Barbara S Frazier
`( 74 ) Attorney , Agent , or Firm Booth Udall Fuller , PLC
`
`( 57 )
`ABSTRACT
`Inorganic anions nitrate and nitrite influence metabolic rate
`and glucose homeostasis . Infusion of nitrite iv caused an
`acute drop in resting energy expenditure ( oxygen consump
`tion ) and nitrate , when given perorally , caused a reduction in
`oxygen consumption during exercise and a depression of the
`increase in blood glucose observed after an oral glucose
`tolerance test . The doses of nitrate and nitrite did not cause
`any detectable change in methemoglobin levels of blood .
`Also , nitrate and nitrite did not alter lactate levels in blood .
`This discovery provides useful treatments to regulate the
`energy expenditure and glucose homeostasis of a mammal
`by administration of inorganic nitrite and / or nitrate .
`
`24 Claims , 13 Drawing Sheets
`
`Systolic
`Diastolic
`MAP
`
`
`
`140-1
`130
`Blood pressure ( mmHg )
`120
`110
`100
`90
`80
`70
`
`8
`
`14
`
`20
`
`Days
`
`Human Power of N Company
`EX1069
`Page 1 of 31
`
`

`

`US 10,842,813 B2
`Page 2
`
`Related U.S. Application Data
`26 , 2008 , now Pat . No. 10,406,118 , application No.
`15 / 966,508 , which is a continuation - in - part of appli
`cation No. 14 / 830,937 , filed on Aug. 20 , 2015 , now
`Pat . No. 10,555,968 , which is a continuation of appli
`cation No. 12 / 528,798 , filed as application No. PCT /
`SE2008 / 050211 on Feb. 26 , 2008 , now Pat . No.
`9,180,140 .
`
`( 60 ) Provisional application No. 60 / 919,709 , filed on Mar.
`22 , 2007 .
`
`( 51 ) Int . Ci .
`A61K 9/08
`A61K 31/05
`A61K 31/198
`A61K 31/375
`A61K 33/14
`A61K 36/185
`A61K 45/06
`A23L 2/02
`A23L 2/52
`A23L 33/105
`A23L 33/135
`A23L 33/16
`A61K 36/21
`A61K 33/30
`A61P 9/12
`COID 9/00
`( 52 ) U.S. CI .
`CPC
`
`( 2006.01 )
`( 2006.01 )
`( 2006.01 )
`( 2006.01 )
`( 2006.01 )
`( 2006.01 )
`( 2006.01 )
`( 2006.01 )
`( 2006.01 )
`( 2016.01 )
`( 2016.01 )
`( 2016.01 )
`( 2006.01 )
`( 2006.01 )
`( 2006.01 )
`( 2006.01 )
`
`A23L 33/135 ( 2016.08 ) ; A23L 33/16
`( 2016.08 ) ; A61K 9/0056 ( 2013.01 ) ; A61K
`970095 ( 2013.01 ) ; A61K 9/08 ( 2013.01 ) ; A61K
`31/05 ( 2013.01 ) ; A61K 31/198 ( 2013.01 ) ;
`A61K 31/375 ( 2013.01 ) ; A61K 33/14
`( 2013.01 ) ; A61K 33/30 ( 2013.01 ) ; A61K
`36/185 ( 2013.01 ) ; A61K 36/21 ( 2013.01 ) ;
`A61K 45/06 ( 2013.01 ) ; A61P 9/12 ( 2018.01 ) ;
`A23V 2002/00 ( 2013.01 ) ; COID 9/00
`( 2013.01 )
`
`( 56 )
`
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`
`Page 2 of 31
`
`

`

`US 10,842,813 B2
`Page 3
`
`( 56 )
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`Bjone et al . , Nitrite in Saliva Increases Gastric and Mucosal Blood
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`Poderoso et al . , Nitric oxide Regulates Oxygen Uptake and Hydro
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`macology 14 ( Suppl . 9 ) : S84-888 , 1989 .
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`Crawford et al . , Hypoxia , Red blood Cells , and Nitrite Regulate
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`Chong , Relationship Between Circulating levels of Nitrates and
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`submitted to fulfill the requirements of the Degree of Master of
`Science to the Chinese University of Hong Kong , 2002 .
`Bjorne , The Nitrite Ion :
`Its Role in Vasoregulation and Host
`Defenses , from the Department of Physiology and Pharmacology
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`Instititutet , Stockholm , Sweden , ( 2005 ) .
`Laycock et al . , Role of Nitric Oxide in the Control of Renal Oxygen
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`Nitrates and Nitrite to Nitric Oxide Under Hypoxic Conditions ,
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`Ceremuzynski et al . , Effect of Supplemental Oral L - Arginine on
`Exercise Capacity in Patients with Stable Angina Pectoris , The
`American Journal of Cardiology , vol . 80 , pp . 331-333 , 1997 .
`Sar et al . , Effects of Combination of Nitrate with 1-4Galacto
`oligosaccharides and Yeast on Methane Emissioni from Sheep ,
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`Bohuslavs Kyi , Effect of Nitric Oxide on the Efficiency of Oxygen
`Consumption by the Working Skeeletal Muscles in Fatigue , Fiziol
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`Gladwyn et al . , Nature Chemical Biology , vol . 1 No. 6 Nov. 2005 ,
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`2006 .
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`“ Effect of dietary nitrate on oxygen cost during
`exercise ” , Acta Physiol ( Oxf ) , 191 ( 1 ) : 59-66 ( Sep. 2007 ) .
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`nitric oxide prduction , metabolism and performance in intermittent
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`and metabolic adaptation in athletes ” , Int J Sport Nutr Exerc Metab ,
`19 : 355-65 ( 2009 ) .
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`“ A Single oral intake of arginine does not affect
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`performance during repeated Wingate anaerobic test ” , J Sports Med
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`1 , 2005 ) , pp . 668-681 , XP0050130963 , ISSN : 0891-5849 , 001 : 10.1
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`* cited by examiner
`
`Page 3 of 31
`
`

`

`U.S. Patent
`
`Nov. 24 , 2020
`
`Sheet 1 of 13
`
`US 10,842,813 B2
`
`Vasodilation Cardioprotection Inhibition of platelets
`
`FIGURE 1
`
`Inactivation of NO
`
`superoxide Free radicals e.g
`
`
`NO
`
`Nitrate Initrite
`
`polyphenols
`
`3 .
`
`1 .
`
`NO synthase
`
`polyphenols )
`
`5 .
`
`4b .
`
`2 .
`
`4a .
`
`Nitros amines
`
`Page 4 of 31
`
`

`

`U.S. Patent
`
`Nov. 24 , 2020
`
`Sheet 2 of 13
`
`US 10,842,813 B2
`
`FIGURE 2
`
`Sodium nitrite infused iv
`
`basal
`
`1
`nmoles / kg / min
`
`10
`
`0.325
`
`0.300
`
`0.275
`VO2 ( ml / min )
`0.250
`
`0.225
`
`0.200
`
`Page 5 of 31
`
`

`

`U.S. Patent
`
`Nov. 24 , 2020
`
`Sheet 3 of 13
`
`US 10,842,813 B2
`
`FIGURE 3
`
`400
`350
`300
`250
`200
`
`
`
`Plasma nitrite ( nM )
`
`150 MI M
`
`100
`50
`0
`
`Rest
`
`Exercise
`
`Rest
`
`Exercise
`
`Page 6 of 31
`
`

`

`U.S. Patent
`
`Nov. 24 , 2020
`
`Sheet 4 of 13
`
`US 10,842,813 B2
`
`FIGURE 4
`
`VO2 NIT
`VO2 CON
`HR NIT
`--- HR CON
`
`5,5
`
`5
`
`4,5
`
`4
`
`3,5
`VO2 ( l / min )
`
`3
`
`2,5
`
`2
`
`1,5
`
`1
`
`45 %
`
`60 %
`
`70 %
`
`80 %
`
`85 %
`
`max
`
`200
`
`180
`
`160
`
`140
`
`120
`
`100 HR ( bpm )
`
`80
`
`60
`
`40
`
`20
`
`0
`
`Page 7 of 31
`
`

`

`U.S. Patent
`
`Nov. 24 , 2020
`
`Sheet 5 of 13
`
`US 10,842,813 B2
`
`FIGURE 5
`
`4.25
`
`4.00
`
`3.75
`VO2 ( l / min )
`3.50
`
`3.25
`
`3.00
`
`Placebo
`
`Nitrate
`
`Page 8 of 31
`
`

`

`U.S. Patent
`
`Nov. 24 , 2020
`
`Sheet 6 of 13
`
`US 10,842,813 B2
`
`FIGURE 6
`
`18
`
`16
`
`14
`
`
`
`Plasma lactate ( mmol / l )
`
`12
`
`10
`
`8
`
`4 I
`
`2
`
`1
`
`T
`
`45 %
`
`60 %
`
`70 %
`
`80 %
`
`85 %
`
`max
`
`Page 9 of 31
`
`

`

`U.S. Patent
`
`Nov. 24 , 2020
`
`Sheet 7 of 13
`
`US 10,842,813 B2
`
`FIGURE 7a
`
`A Nitrate
`Placebo
`
`0
`
`10 30 45 60 75 90 120
`Time
`
`9
`8
`7 -
`6 T
`5
`B - Glucose 4
`
`3
`2
`1
`0
`
`Page 10 of 31
`
`

`

`U.S. Patent
`
`Nov. 24 , 2020
`
`Sheet 8 of 13
`
`US 10,842,813 B2
`
`FIGURE 7b
`
`Placebo
`Nitrate
`
`0
`
`10
`
`30
`
`60
`45
`Time
`
`75
`
`90
`
`120
`
`8
`
`7
`
`6
`
`B - Glucose 4
`
`3
`
`2
`
`1
`
`0
`
`Page 11 of 31
`
`

`

`U.S. Patent
`
`Nov. 24 , 2020
`
`Sheet 9 of 13
`
`US 10,842,813 B2
`
`9
`8
`7
`
`UO
`B - Glucose 3
`
`2
`1
`0
`
`FIGURE 7c
`
`Nitrate
`Placebo
`
`0
`
`10
`
`30 45 60 75 90 120
`Time
`
`Page 12 of 31
`
`

`

`U.S. Patent
`
`Nov. 24 , 2020
`
`Sheet 10 of 13
`
`US 10,842,813 B2
`
`FIGURE 8
`
`87
`
`B - glucose
`
`7
`
`6
`
`5
`
`4
`
`T
`
`-10
`
`0
`
`15
`
`30
`
`45
`
`60 75
`
`20
`
`105
`
`120 ( min )
`
`Page 13 of 31
`
`

`

`U.S. Patent
`
`Nov. 24 , 2020
`
`Sheet 11 of 13
`
`US 10,842,813 B2
`
`FIGURE 9
`
`beetroot juice
`3-4 dl daily
`
`Systolic
`Diastolic
`MAP
`
`4
`
`6
`
`7
`Days
`
`00
`
`14
`
`20
`
`140
`130
`Blood pressure ( mmHg ) 110
`120
`
`
`
`100
`90
`
`80 50 70
`
`Page 14 of 31
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`

`

`U.S. Patent
`
`Nov. 24 , 2020
`
`Sheet 12 of 13
`
`US 10,842,813 B2
`
`FIGURE 10
`
`O Nitrate
`Nitrate + Allopurinol
`Placebo
`
`b
`
`1500
`
`1200
`
`O Nitrate
`A Nitrate + Allopurinol
`
`
`
`Plasma Nitrite ( nM )
`
`900
`
`a
`
`10007
`
`800
`
`
`
`Plasma Nitrate ( UM )
`
`600
`
`p = 06
`
`M : 1
`
`400
`
`200
`
`-30
`
`0
`
`?
`
`60
`30
`15
`Time ( min )
`
`90 120
`
`3000
`
`2000
`
`
`
`Plasma Nitrite ( nM )
`
`600
`
`300
`
`T
`
`so
`
`60
`
`120
`
`- 30
`
`1
`
`30
`15
`Time ( min )
`Placebo
`Nitrate 10mg / kg
`
`T
`
`1000 -
`
`C57BL / 6
`
`Germ Free
`
`eNOS-
`
`Page 15 of 31
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`

`U.S. Patent
`
`Nov. 24 , 2020
`
`Sheet 13 of 13
`
`US 10,842,813 B2
`
`FIGURE 11
`
`400
`
`300
`
`ICH
`
`
`
`% Change in blood flow 200
`
`HE HAH
`
`100
`
`???
`
`0
`
`0 1 2 3
`
`5
`
`7
`
`9 10
`
`- Nitrate
`Placebo
`
`?
`
`? ? ? ?
`
`? ?
`
`? ? ?
`
`20 30 40
`Time
`
`50 60
`
`Page 16 of 31
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`

`

`US 10,842,813 B2
`
`1
`COMPOSITIONS OF NITRATES AND
`METHODS OF USE THEREOF
`
`2
`dietary nitrate in humans . Gut . 1997 , no . 40 , p . 211-214 ; and
`Weitzberg , 1998 , supra ) is absorbed from the circulation by
`the salivary glands , secreted in saliva and partly converted
`to nitrite in the oral cavity by nitrate reducing bacteria .
`RELATED APPLICATIONS
`5 Swallowed nitrite can then enter the systemic circulation .
`Indeed , a recent study shows that ingestion of nitrate results
`This application is a continuation application of earlier
`in a sustained increase in circulating nitrite levels ( LUND
`U.S. Utility Patent Application to Jon Lundberg and Eddie
`Weitzberg , entitled “ Use of Nitrites and Nitrates and Com BERG , JO , et al . Inorganic nitrate is a possible source for
`positions Containing These , " application Ser . No. 12/528 ,
`systemic generation of nitric oxide . Free Rad Bio Med .
`794 , filed Jun . 17 , 2013 , now pending , which is a national 10 2004 , vol . 37 , no . 3 , p . 395-400 ) . Further reduction of nitrite
`stage application of PCT application No. PCT / SE08 / 50212 ,
`into bioactive NO can occur spontaneously in acidic or
`filed Feb. 26 , 2008 , which claims the benefit of the filing
`reducing environments ( Benjamin et al . 1994 , supra , Lun
`date of U.S. Provisional Patent Application 60 / 919,709 to
`dberg et al . 1994 , supra ) but is also greatly enhanced by
`Jon Lundberg and Eddie Weitzberg , filed on Mar. 22 , 2007 ,
`the disclosures of all of which being hereby incorporated 15 various proteins and enzymes including deoxyhemoglobin
`in blood ( COSBY , K , et al . Nitrite reduction to nitric oxide
`entirely herein by reference .
`by deoxyhemoglobin vasodilates the human circulation . Nat
`This application is also a continuation application of
`Med . 2003 , vol . 9 , no . 12 , p . 1498-505 ) , deoxymyoglobin
`earlier U.S. Utility Patent Application to Jon Lundberg and
`( SHIVA , S. et al . Deoxymyoglobin is a Nitrite Reductase
`Eddie Weitzberg , entitled “ Performance Enhancing Compo
`sitions and Use Thereof , ” U.S. patent application Ser . No. 20 That Generates Nitric Oxide and Regulates Mitochondrial
`Respiration . Circ Res . 9 Feb. 2007 ) , xanthine oxidase ( MIL
`14 / 830,937 , filed Aug. 20 , 2015 , now pending , which is a
`continuation application of U.S. patent application Ser . No.
`LAR , T M , et al . Xanthine oxidoreductase catalyses the
`12 / 528,798 , filed Aug. 26 , 2009 , now issued as U.S. Pat . No.
`reduction of nitrates and nitrite to nitric oxide under hypoxic
`conditions . FEBS Lett . 8 May 1998 , vol . 427 , no . 2 , p .
`9,180,140 , which was a national stage application of PCT
`application No. PCT / SE 2008/050211 , filed Feb. 26 , 2008 , 25 225-8 ) and possibly by enzymes of the mitochondrial respi
`ratory chain ( for review see LUNDBERG , JO , et al . Nitrate ,
`which is a claims priority to Swedish Patent Application
`0700520-0 , which was filed Feb. 26 , 2007 and Swedish
`bacteria and human health . Nat Rev Microbiol . 2004 , vol . 2 ,
`no . 7 , p . 593-602 ; LUNDBERG , J O , et al . NO generation
`Patent Application 0700729-0 , which was filed Mar. 22 ,
`2007 , the disclosures of all of which being hereby incorpo
`from nitrite and its role in vascular control . Arterioscler
`30 Thromb Vasc Biol . 2005 , vol . 25 , no . 5 , p . 915-22 ; and
`rated entirely herein by reference .
`GLADWIN , M T , et al . The emerging biology of the nitrite
`anion . Nat Chem . Biol . 2005 , vol .
`1 , no . 6 , p . 308-14 ) .
`FIELD OF THE INVENTION
`NOS - independent NO production seems to complement the
`endogenous NO production especially during ischemia and
`The present invention relates to the field of performance
`enhancing nutritional foods and food supplements , liquid 35 acidosis when oxygen availability is low and the NO syn
`and solid edible products such as sport drinks , energy drinks
`thases operate poorly ( Zweier et al . 1995 , supra ; Weitzberg
`and energy bars . The present invention also relates to the
`et al , 1998 , supra ; DURANSKI , M R , et al . Cytoprotective
`field of medicine and pharmaceuticals , in particular phar
`effects of nitrite during in vivo ischemia - reperfusion of the
`maceuticals and therapeutic methods for lowering metabolic
`heart and liver . J Clin Invest . 2005 , vol . 115 , no . 5 , p .
`rate , oxygen consumption and / or glucose homeostasis in a 40 1232-40 ; Lundberg et al , 2004 , supra ) . Tissue acidosis and
`human patient or another mammal , based on the adminis
`relative hypoxia is present also during physical exercise and
`tration of nitrates and / or nitrites to said patient or mammal .
`in this metabolic state , bioactivation of nitrite is likely
`enhanced .
`Recent studies indicate that nitrate and nitrite can have
`BACKGROUND OF THE INVENTION
`45 significant biological effects in the body and that these
`effects may be beneficial ( LUNDBERG , Jon O. , et al .
`NO is involved in control of cellular respiration through
`Nitrate , becteria and human health . Nat Rev Microbiol .
`interaction with enzymes of the mitochondrial respiratory
`chain ( for review see MONCADA , S , et al . Does nitric oxide
`2004 , no . 2 , p . 593-602 ) . For example , the nitrite anion can
`modulate mitochondrial energy generation and apoptosis ?.
`cause vasodilatation at near physiological concentrations
`Nat Rev Mol Cell Biol . 2002 , vol . 3 , no . 3 , p . 214-20 ) . The 50 when tested in vitro ( MODIN , A. , et al . Nitrite - derived nitric
`classical means by which NO production occurs is the
`oxide : a possible mediator of ‘ acidic - metabolic ' vasodila
`L - arginine pathway , where NO is synthesized by specific
`tion . Acta Physiol Scand . 2001 , vol . 171 , p . 9-16 ) or when
`enzymes , the NO - synthases . A fundamentally different alter
`infused intra - arterially to humans ( COSBY , K. , et al . Nitrite
`native way of generating NO has been described more
`reduction to nitric oxide by deoxyhemoglobin vasodilates
`recently ( LUNDBERG , J O , et al . Intragastric nitric oxide 55 the human circulation . Nat Med . 2003 , no . 9 , p . 1498-505 ) .
`production in humans : measurements in expelled air . Gut .
`Nitrate can be converted to nitrite in vivo in a process
`1994 , vol . 35 , no . 11 , p . 1543-6 ; BENJAMIN , N , et al .
`dependent on commensal bacteria ( SPIEGELHALDER , B. ,
`Stomach NO synthesis . Nature . 7 Apr. 1994 , vol . 368 , no .
`et al . Influence of dietary nitrate on nitrite content of human
`6471 , p . 502 ; ZWEIER , J L , et al . Enzyme - independent
`saliva : possible relevance to in vivo formation of N - nitroso
`formation of nitric oxide in biological tissues . Nat Med . 60 compounds . Food Cosmet Toxicol . 1976 , no . 14 , p . 545
`1995 , vol . 1 , no . 8 , p . 804-9 ; and WEITZBERG , E , et al .
`548 ) . When nitrate is ingested it is rapidly absorbed into
`Nonenzymatic nitric oxide production in humans . NO Biol .
`blood and then accumulates in saliva . In the oral cavity
`Chem . 1998 , no . 2 , p . 1-7 ) . In this NOS - independent path
`bacteria reduce parts of the dietary nitrate to nitrite and
`way the inorganic anions nitrate ( NO3- ) and nitrite ( NO
`nitrite can then enter the systemic circulation . ( LUND
`.sub.2- ) are reduced in vivo to form NO . Dietary nitrate 65 BERG , Jon O. , et al . Inorganic nitrate is a possible source for
`( found mainly in green leafy vegetables ) ( MCKNIGHT , G systemic generation of nitric oxide . Free Radic Biol Med .
`M. Chemical synthesis of nitric oxide in the stomach from
`2004 , vol . 37 , p . 395-400 ) .
`
`Page 17 of 31
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`

`US 10,842,813 B2
`
`4
`3
`effects of administering exogenous NO on exercise are
`In vitro studies published in the 1990s show that NO is a
`largely unknown . In addition , studies in healthy humans are
`modulator of mitochondrial respiration via reversible inhi
`bition of cytochrome c oxidase ( CARR , G J. et al . Nitric
`scarce .
`Interestingly , the marketing of some currently available
`oxide formed by nitrite reductase of Paracoccus denitrificans
`is sufficiently stable to inhibit cytochrome oxidase activity 5 food supplements for athletes and bodybuilders refer to the
`and is reduced by its reductase under aerobic conditions .
`vasodilatory effect of NO . One example is “ NOX2 ”
`( Bodyonics , Ltd. , USA ) , a product said to contain arginine
`Biochim Biophys Acta . 15 May 1990 , vol . 1017 , no . 1 , p .
`57-62 ; BOLANOS , J P , et al . Nitric oxide - mediated inhibi
`alpha - ketoglutarate ( A - AKG ) and arginine - ketoisocaproate
`( A - KIC ) and allegedly capable of boosting short term nitric
`tion of the mitochondrial respiratory chain in cultured astro
`cytes . J. Neurochem . 1994 , vol . 63 , no . 2 , p . 910-6 ; 10 oxide levels . Other products contain L - arginine , from which
`NO is synthesized by the NOS enzymes , and the beneficial
`BROWN , G C , et al . Nanomolar concentrations of nitric
`effects of NO are often referred to , however without offering
`oxide reversibly inhibit synaptosomal respiration by com
`more detailed explanations .
`peting with oxygen at cytochrome oxidase . FEBS Lett . 19
`The relation between peak work rate and resting levels of
`Dec. 1994 , vol . 356 , no . 2-3 , p . 295-8 ; CLEETER , M W , et 15 nitrate in plasma and urine from subjects with different
`al . Reversible inhibition of cytochrome c oxidase , the ter
`levels of physical fitness has been studied ( Jungersten et al . ,
`minal enzyme of the mitochondrial respiratory chain , by
`Both physical fitness and acute exercise regulate nitric oxide
`nitric oxide . Implications for neurodegenerative diseases .
`formation in healthy humans . J Appl Physiol 82 : 760-764 ,
`FEBS Lett . 23 May 1994 , vol . 345 , no . 1 , p . 50-4 ; and
`1997 ) . A positive relationship between physical fitness and
`SCHWEIZER , M , et al . Nitric oxide potently and reversibly 20 formation of NO at rest was found and it was hypothesised
`deenergizes mitochondria at low oxygen tension . Biochem
`that this positive relationship helps to explain the beneficial
`Biophys Res Comm . 1994 , no . 204 , p . 169-75 ) . NO may also
`effects of physical exercise on cardiovascular health . In
`interact at other sites of the mitochondrial respiratory chain
`Jungersten's study nitrate was used solely as a marker of NO
`and in the Krebs cycle ( for review see Moncada , supra ) .
`production and the authors state several times that nitrate is
`While this important action of NO has been very well 25 a stable and inert end product of NO and that it is biologi
`characterised in cell cultures , less is known about its physi
`cally inactive .
`ological relevance in vivo . To date the focus among
`Nitrate ( NO3- ) and nitrite ( NO2 ) are generally viewed as
`researchers has been on the cardiovascular effects of nitrite
`unwanted residues in the food chain with potentially harmful
`after its in vivo reduction to the vasodilator nitric oxide ( NO )
`effects ( Joint FAO / WHO Expert Committee on Food Addi
`( COSBY et al . ( supra ) ; DURANSKI , M. R. , et al . Cytopro- 30 tives ( JECFA ) . Safety Evaluation of Certain Food Additives .
`tective effects of nitrite during in vivo ischemia - reperfusion
`WHO , 1970. ISBN 9241660503 ; TANNENBAUM , S. R. , et
`of the heart and liver . J Clin Invest . 2005 , vol . 115 , p .
`al . Nitrite in human saliva . Its possible relationship to
`1232-1240 ; GLADWIN , M. T. ,
`al . The emerging biology
`nitrosamine formation . J cancer Ins . 1974 , vol . 53 , p . 79-84 ;
`of the nitrite anion . Nat Chem Biol . 2005 , no . 1 , p . 308-14 ;
`BARTSCH , H. , et al . Inhibitors of endogenous nitrosation :
`LARSEN , F. J. , et al . Effects of dietary nitrate on blood 35 mechanisms and implications in human cancer prevention .
`pressure in healthy volunteers . N Engl J Med . 2006 , vol .
`Mutation Res . 1988 , vol . 202 , p . 307-324 ) . Proposed harm
`355 , p . 2792-3 ) . WO 2005/004884 A ( US GOVERNMENT
`ful effects of these anions include promotion of gastric
`ET AL . ) Jan. 20 , 2005 and WO 2005/007173 A ( US GOV cancers and other malignancies and development of methe
`ERNMENT ET AL . ) Jan. 27 , 2005 describe a method to
`moglobinemia in infants . Because of this the levels of
`administer a nitrite salt specifically to obtain vasodilatation 40 nitrate / nitrite are strictly regulated in food and drinking
`in a subject . No effects of low - dose nitrate / nitrite on energy
`water .
`expenditure or glucose homeostasis or the effects of NO on
`cellular respiration during phy