Redox Diagnostics

01/07/2025

One of the most popular trends in healthcare these days is vitamin infusion therapy, where a mix of vitamins, minerals and antioxidants (such as glutathione), is administered intravenously to patients.

The claims are very bold, ranging from detoxification of the body to regulation of sleep, mood and appetite and an overall superior hydration. They start from the premise that the mix of vitamins and antioxidants provide an antioxidant protection that is responsible for the plethora of benefits mentioned in the advertising.

But how can a healthcare professional claim that a mix of antioxidants delivered in bulk via intravenous delivery is good for the patient without an objective assessment of the redox status of the patient? How does the healthcare professional know what is the right dose for any patient? Or how much is too much? Is the actual ‘one size fits all’ approach the right approach?

We contend it is not and here is why:

- redox homeostasis is the most delicate and important control mechanism in a living system, given its critical role in maintaining the functional integrity of all its other components.
- a redox system is made of two major and complementary components: the pro-oxidant component and the reductive (or antioxidant) component. Redox homeostasis in the system is achieved when the two components are in balance.
- an imbalance created due to either an increase in the pro-oxidant components or a deficiency in reductive components leads to oxidative stress, a condition known to be at the root of any acute and chronic diseases.
- on the other hand, a decrease of the pro-oxidant components and/or an increase of the reductive components leads to a redox imbalance called reductive stress, an equally detrimental condition, less mentioned and explored in scientific literature.
- administration of reductive (antioxidants) substrates such as vitamin C and glutathione without measurement of patients’ redox status comports risks, since both substrates become pro-oxidants after consuming their reductive action and by reducing transition metals in an oxidized state such as Fe3+ and Cu 2+, thus initiating an oxidative chain of reactions. This very relevant scientific paper highlights the premise that IV administration of vitamin C leads to oxidative inactivation of proteins.
https://www.sciencedirect.com/science/article/abs/pii/S0002916523319828

There is a simple and effective solution to finding patients’ Redox Status in minutes, at Point-of-Care:

- FRAS5 – dedicated spectrophotometer with integrated centrifuge for in-minutes determination of redox status in plasma and saliva samples
- d-ROMs fast test – determines the pro-oxidant status in plasma samples
- PAT test – determines the antioxidant status in plasma samples
- SAT test – determines the antioxidant status in saliva samples
- Oxidative Stress Index (OSI) – an integrated index calculated from the values of d-ROMs fast test and PAT test, that allows for a simpler interpretation and communication of patients’ redox status

The device and tests are validated by over 1,800 scientific papers published worldwide.

More info here: https://redoxdiagnostics.com/fras5-2/

04/03/2024

Today is an important moment for our company as we change the name and internet domain name under which we operate from Innovatics Laboratories, Inc. to Redox Diagnostics, Inc. and www.redoxdiagnostics.com, so that it better reflects the nature of our activities. It reaffirms our commitment to bring to the market the most advanced solutions for the assessment of redox status in biological systems.

After years of research in the field of redox biology and related disciplines, we feel compelled to share the knowledge with like-minded individuals who understand and are willing to leverage on the major shift currently underway in the healthcare space.

Our commercial endeavors will be matched by our educational efforts, aiming to contribute to the advancement of diagnostic and therapeutic solutions rooted in the right understanding of physiological and pathophysiological processes.

01/18/2024

HYDROGEN PEROXIDE - A CRITICALLY IMPORTANT MOLECULE

As noted previously, hydrogen peroxide is one of the Reactive Oxygen Species (ROS), along with hydroxyl radical (HO·), peroxyl radical (HOO·) and superoxide anion (O-2), with a moderate oxidation power.
Despite its simplicity and relative instability, hydrogen peroxide is a versatile molecule that plays pivotal roles in multiple biological processes; therefore, its synthesis and degradation are both tightly regulated processes.

I. Hydrogen peroxide synthesis
There are multiple pathways of hydrogen peroxide synthesis, such as:
a. Dismutation of superoxide anion (O-2) by superoxide dismutase (SOD), per following reaction:

2H+ + 2O-2 -> O2 + H2O2

b. Degradation of monoamine neurotransmitters: hydrogen peroxide is a catalytic reaction product from the mitochondrial outer membrane enzymes monoamine oxidases (MAO) A and B. For example, oxidative deamination of dopamine (DA) by monoamine oxidase (MAO) produces hydrogen peroxide (H2O2)
and the reactive aldehyde DOPAL (3,4 dihydroxyphenilacetaldehyde).

c. Byproduct of fatty acids hydrolysis in the peroxisomes

Peroxisomes are oxidative organelles, whose main metabolic function is the breakdown of very long chain fatty acids through beta-oxidation. In presence of oxygen (O2), the long chain fatty acids are converted to medium chain fatty acids, that are then transported to the mitochondria for further breakdown to water and carbon dioxide.
Oxidation of long fatty acids leads to formation of hydrogen peroxide per following reaction:

R-H2 + O2 -> R + H2O2

d. During oxidative protein folding in the endoplasmic reticulum.
Oxidative protein folding in the endoplasmic reticulum (ER) is a significant source of hydrogen peroxide (H2O2). This process
involves protein disulfide isomerase (PDI) working in concert with ER oxidoreductin 1 (Ero1) to catalyze the formation of disulfide bonds. Molecular oxygen is the ultimate electron acceptor in this process and yields one H2O2molecule for every disulfide bond formed. Excessive amounts of hydrogen peroxide are transported out of the ER through the transmembrane channels Aquaporins-11 into the cytosol for degradation by glutathione (GSH).

II. Hydrogen peroxide degradation
There are several pathways of hydrogen peroxide degradation, such as:
a. In peroxisomes, hydrogen peroxide is used to oxidize other substrates in a reaction mediated by the enzyme catalase:

R-H2 + H2O2 -> R + 2H2O

If hydrogen peroxide accumulates in excess, catalase will degrade it to oxygen and water, per following reaction:

2H2O2 -> O2 + H2O

To be noted that catalase is an enzyme present only in the peroxisomes and not in the cytosol and the mitochondria.

b. In cytosol and mitochondria, excessive amounts of hydrogen peroxide are degraded by reduced glutathione (GSH) to water and oxidized glutathione (GS-SG), in a reaction catalyzed by glutathione peroxidase (GPx):
H2O2 + 2GSH -> 2H2O + GS-SG

c. In the erythrocytes and mitochondria, hydrogen peroxide is also degraded by peroxiredoxin (Prx), with formation of sulfenic acid and water:
PrxSH + H2O2 -> PrxSOH + H2O

d. In the presence of photons, hydrogen peroxide undergoes a spontaneous disproportionation to water and oxygen, per following reaction:
2H2O2 -> O2 + H2O

III. Hydrogen peroxide biological functions
Hydrogen peroxide is involved in multiple critical biological processes, such as:
a. Immune response – hydrogen peroxide is synthesized by the macrophages in response to invading pathogenic bacteria and is a critical part in the bacterial degradation through oxidative degradation of its organic substrates.
It also acts as a substrate for hypochlorous acid (HOCl) formation in the neutrophils, as part of the innate immune response, in a reaction mediated by myeloperoxidase (MPO).

b. Energy metabolism
As noted in a previous post, it is our understanding that generation of hydroxyl radical (HO·) from hydrogen peroxide by Fenton chemistry is part of a novel pathway of energy production that involves a HOMO/LUMO reaction between hydroxyl radical (HO·) and reduced glutathione (GSH, as follows:

H2O2 + Fe2+  HO· + HO- + Fe3+
2 HO· + 2 GSH  2 H20 + GS-SG + 2 photons

c. Signaling
Oxidation of the thiol group (-SH) of the methionine residue (Met-SH) to methionine sulfoxide (Met-S=O) is a signal that triggers protein phosphorylation, while reduction of the methionine sulfoxide is a signal for dephosphorylation. This process has major implications in proteins functionality.

d. Modulation of redox sensors
It is well established that reversible oxidative post-translational modifications of the cysteine residues by hydrogen peroxide represent an important mechanism that regulates protein structure and function. For example, formation of disulfide bonds between ATP synthase redox sensors from alpha subunit Cys294 and gamma subunit Cys103 is correlated with inhibition of ATP synthesis and ATP hydrolysis. Reversal of disulfide bond at Cys294 results in the recovery of ATP synthase activity.

It is apparent that in small amounts and under tight control of synthesis and degradation, hydrogen peroxide plays key roles in a multitude of physiological processes. However, accumulation of excessive amounts of hydrogen peroxide it is very damaging and is associated with the development of many chronic and degenerative conditions. We will address this topic in the next post.
Given the dual nature of hydrogen peroxide as a benefactor in small, physiological amounts and a detrimental molecule in excessive amounts, monitoring of the level of peroxides in blood samples should be a top priority in any healthcare protocol. This can be easily done in-office or in laboratory by using d-ROMs test and the dedicated instrument FRAS5. More information here https://innovaticslabs.com/d-roms-fast-test/

The d-ROMs fast test, developed initially by the renowned scientist Mauro Carratelli and upgraded by H&D srl, is a photometric test that allows to assess the pro-oxidant status in a biological sample, by measuring the concentration of hydroperoxides (ROOH).

12/12/2023

PEROXIDES FORMATION AND THEIR SIGNIFICANCE

We mentioned in a previous post that hydroxyl radical (HO·) formation is pivotal for energy production by reacting with glutathione (GSH) in a HOMO/LUMO reaction.

What happens if there is an imbalance between hydroxyl radical formation and reduced glutathione availability?

In absence of enough glutathione supply, hydroxyl radical (HO·), a very reactive molecule, will seek to stabilize by oxidizing organic substrates with lower reductive power, such as lipids, proteins and nucleic acids and initiates a chain reaction that leads to peroxides formation in the early stage and to other oxidation byproducts in the more advanced oxidation stages. We will discuss here the process of lipid peroxidation.
The lipid peroxidation chain reaction happens in three steps: Initiation, Propagation, Termination.

1. Initiation.
In the initiation step the hydroxyl radical (HO·) extracts the hydride ion (H-) required for stabilization from a lipid (LH), leading to formation of a water molecule (H20) and a carbon-centered lipid radical, per following reaction:

HO· + LH -> H20 + L·

2. Propagation
The newly formed lipid radical (L·) reacts with an oxygen molecule and generates a lipid peroxyl radical, per following reaction:

L· + O2 -> LOO·

The peroxyl radical (LOO·) is seeking to stabilize as well by reacting with another lipid molecule (LH), generating a more stable molecule of lipid hydroperoxide (LOOH) and another lipid radical (L·), per following reaction:

LOO· + LH -> LOOH + L·

The newly formed lipid radical (L·) further undergoes the reaction with oxygen (O2) generating a peroxyl radical (LOO·) and later another lipid radical (L·); the reaction propagates until a reductive substrate terminates it.

3. Termination
In presence of reductive substrates, such as glutathione (GSH), Vitamin C, vitamin E, the oxidation chain reaction is interrupted by stabilizing the reactive species through multiple mechanisms:

a. Lipid peroxyl radicals (L-OO·) are reduced to more stable lipid hydroperoxides (L-OOH) by vitamin C and vitamin E.

b. Lipid hydroperoxides are reduced by glutathione (GSH) to more stable and less toxic lipid alcohols or hydroxy fatty acids, in a reaction mediated by glutathione peroxidase 4 (GPX4):

L-OOH + 2GSH -> L-OH + GS-SG + H2O

c. Lipid radicals (L·) are reduced to stable lipid molecules (LH) by glutathione (GSH) in a reaction mediated by glutathione peroxidase:

2L· + 2GSH -> 2LH + GS-SG

In absence of reductive substrates, the lipid peroxides (LOOH) undergo further oxidation into more advanced oxidation byproducts, such as malondialdehyde (MDA) and 4-hydroxynonenal (HNE).
What is the significance of hydroperoxides formation and why it is important to monitor the level of hydroperoxides in blood or plasma samples?

Per the explanation above, hydroperoxides are the first line of oxidation byproducts and therefore the EARLIEST WITNESS of oxidative stress - an imbalance between the Reactive Oxygen Species (ROS) formation and the reductive capacity of the endogenous antioxidant system. The implications are multiple, depending on the organic substrates affected by the oxidative damage, from impairment of energy production to membrane destabilization and permeabilization due to lipid peroxidation, to mutagenic effects due to oxidative damage of the nucleic acids.

Measurement of the level of hydroperoxides in plasma samples can be done in-office by using our very precise d-ROMs test (determination of Reactive Oxygen Metabolites) and the dedicated analytical instrument FRAS5 (Free Radical Analytical System). It is a very simple and fast procedure that requires a small amount of capillary blood, minimal processing and delivers the results in minutes.
More information available here https://innovaticslabs.com/fras5-2/

Innovatics introduces a convenient and technologically advanced system for the global assessment of oxidative stress, consisting of an innovative Free Radical Analytical System, FRAS5, two blood tests, d-ROMs fast test and PAT test, and one saliva test, SAT test.

12/12/2023

Innovatics Laboratories is a premier supplier of a unique Point-of-Care REDOX testing system, consisting of:

• FRAS5 (Free Radical Analytical System) – dedicated spectrophotometer with centrifuge and printer incorporated, suitable for in-office use.

• two blood tests:
- d-ROMs fast test (determination of Reactive Oxygen Metabolites) – measures the pro-oxidant status in plasma samples.
- PAT test (Plasma Antioxidant capacity Test) – measures the antioxidant/reductive capacity in plasma samples.

• one saliva test:
-SAT test (Saliva Antioxidant capacity Test) – measures the antioxidant/reductive capacity in saliva samples.

• Oxidative Stress Index (OSI) – index that integrates in a single value the information provided by the d-ROMs fast and PAT tests, to simplify the interpretation of REDOX status in plasma samples.

The system provides real-time, accurate assessment of the REDOX status in the tested subjects, which provides valuable information with major implications in the healthcare professionals' decision-making process.

The REDOX status of a subject is indicative of a state of REDOX homeostasis, oxidative stress, or reductive stress. REDOX homeostasis is a state of balance between the production of Reactive Oxygen Species (ROS) and the reductive capacity of the endogenous antioxidant system, resulting in an optimal state of cellular energy production and structural integrity. Oxidative stress is a state of imbalance between increased production of Reactive Oxygen Species (ROS) and reduced reductive capacity of the endogenous antioxidant system. Conversely, reductive stress is a state of imbalance between increased reductive capacity of the endogenous and exogenous antioxidant systems and a reduced production of Reactive Oxygen Species (ROS). Both states of oxidative stress and reductive stress are dysfunctional and, if left uncorrected, may lead to a multitude of acute and chronic pathological conditions.

Oxidative stress is associated with alteration of mitochondrial ATP production, oxidative DNA damage and mutagenesis, cell depolarization, and iron dysregulation.

To maximize clinical outcomes, REDOX homeostasis is the desired REDOX state in any subjects undergoing a medical or dental procedure, particularly surgical procedures; therefore, real-time monitoring of the REDOX status by means of d-ROMs fast and PAT tests and the integrated Oxidative Stress Index (OSI) are of critical importance. The information provided enables the healthcare practitioners to adjust the therapeutic protocols in the preparation phase in order to achieve a state of REDOX homeostasis, and also plays a predictive role of the clinical outcome.

11/07/2023

Hydroxyl radical (HO·) and Glutathione (GSH) – the actual powerhouse of the mitochondria

The current understanding regarding generation and roles of Reactive Oxygen Species (ROS) is multifaceted, having roles attributed as signaling molecules and key players in the immune response and the inflammatory process. They are also largely considered unwanted side effects of the energetic metabolism.
By contrary, our research indicates that in case of their roles in the energetic metabolism, the generation of ROS is quintessential to energy production and not an unwanted side effect. It challenges the actual understanding that energy production takes place at Complex V, where ATP synthesis takes place.
In our hypothesis, there is a coordinated chain of reactions involving generation of Reactive Oxygen Species (ROS) that leads to energy production, as follows:

1. Superoxide anion (O2-) is generated per following reaction, mediated by NADPH oxidase:

2O2 + NADPH -> 2O2- + NADP+ + H+

2. Superoxide anion reacts with two protons (H+) and generates hydrogen peroxide (H2O2) in a reaction mediated by superoxide dismutase (SOD):
2O2- + 2 H+ -> O2 + H2O2

3. In presence of ferrous iron (Fe2+), hydrogen peroxide undergoes the Fenton reaction and generates one hydroxy radical (HO·) and one hydroxyl anion (HO-):

H2O2 + Fe2+ -> HO· + HO- + Fe3+

4. Two hydroxyl radicals (HO·) react with two molecules of glutathione (GSH) and generate two molecules of water, glutathione disulfide and two photons. This is the reaction that generates energy in the mitochondria:

2 HO· + 2 GSH -> 2 H20 + GS-SG + 2 photons

Why are photons generated in the reaction between hydroxyl radical and glutathione?

According to our hypothesis, glutathione and hydroxyl radical undergo a HOMO/LUMO reaction, in which the thiol group (-SH) of glutathione donates a hydride ion (H-) that carries a high energy electron situated on the Highest Occupied Molecular Orbital (HOMO), that will lose energy and, on the way, emit a photon, so that it can fill the Lowest Unoccupied Molecular Orbital (LUMO) of the hydroxyl radical.

The photons emitted are then stored in the phosphorus atoms of the iPO4 present in the mitochondrial matrix, which then moves to Complex V (ATP synthase) to synthesize ATP by joining a molecule of ADP.

5. Oxidized glutathione (GS-SG) is reduced back to two molecules of glutathione by NADPH, in a reaction mediated by glutathione reductase (GSR):
GS-SG – 2 NADPH -> 2 GSH + NADP+

6. NADP+ is reduced back to NADPH in the Pentose Phosphate Pathway, in a reaction mediated by glucose-6-phosphate dehydrogenase, a rate-limiting enzyme.

In conclusion, the high reactivity of hydroxyl radical must be seen as a requirement for energy production in the context of its HOMO/LUMO reaction with glutathione, where one photon is generated by energy cessation by the high energy electron carried out by the thiol group of glutathione, while trying to reach the energy level required to fill the lowest unoccupied molecular orbital of the hydroxyl radical.
Glutathione has its source of hydride ion (H-) that carries the high energy electron from NADPH, which in turn carries it away from a molecule of glucose. So, the ultimate source of energy is the molecule of glucose, but how energy is generated is different in our model compared to the conventional one.

10/30/2023

After many years of research and data collection and integration from multiple scientific disciplines, we believe we arrived at an understanding of the redox processes that take place in biological systems that challenges to a large extent the current understanding of the phenomenon and at the same time offers a new perspective of it, with far reaching implications. As it is clear that the current frame of reference in healthcare does not offer the right solutions, we aim to present the information in a new, different frame of reference that will enable the development of the right therapeutic approaches.

The awareness of the central role oxidative and reductive stress play in the pathophysiology of many acute and chronic and degenerative diseases is quite high in the naturopathic and alternative medicine sectors, but it is still very limited in mainstream medicine and dentistry. We believe this awareness must reach mainstream levels and we aim to contribute to that goal.

While targeted therapies for redox homeostasis are limited, currently, to combinations of natural antioxidant extracts and vitamins, there is very limited testing of the level of oxidative stress in the subjects, leading to high risk of converting oxidative stress to reductive stress, among others.

There is high value in knowing the levels of pro-oxidant and anti-oxidant status in subjects receiving antioxidant therapies and not only. It empowers the healthcare professionals to adjust and maximize the outputs of their therapeutic protocols.
We will start sharing our accumulated knowledge and insights in a series of posts on the company page and will correlate the information with the importance of testing for oxidative stress in general and with our unique FRAS5 testing system in particular.