A Push-Catch System That Enables Effective Detoxification


by Christopher Shade, PhD, and Carrie Decker, ND

 

The human body is exposed to environmental toxins every day from a wide array of sources: particulate matter and diesel fumes in the air,(1,2) heavy metals and other contaminants in the water,(3,4) pesticide and herbicide residues found on foods,(5,6) and even substances like bisphenol A (BPA) via contact with the skin.(7) On a continuous basis, the body must work to eliminate toxic substances that are taken in. If intake exceeds removal, the toxins accumulate within the tissues and cells. These toxins tax our antioxidant systems, which must be upregulated in attempts to reduce cellular damage and death.(8) But even with upregulation, the antioxidant protection system is often depleted by repeated insults and toxin exposure, and this depletion may contribute to disease processes.(9) Chronic exposure to environmental toxins and toxic heavy metals is associated with the development of many types of cancer, respiratory disease, cardiovascular disease, diabetes, infertility, allergies, autoimmune disease, and many other conditions.(10,11,12,13)

Detoxification is the process by which the body eliminates substances that are both endogenous (such as hormones) and exogenous (such as medications, pollutants, metals, and other substances). In addition to cleaning up the diet and eliminating sources of toxicity, the use of chelating substances, antioxidant support, and therapeutic sweating are often the mainstays of detoxification protocols. However, these basic strategies, despite being crucial, lack consideration for other factors such as chronic infections, cholestasis, and enterohepatic recircu-lation of toxic substances that significantly impair the body's ability to detoxify. To comprehensively support detoxification, one must consider and address not only the glutathione system and its enzymes; liver, kidney, and gastrointestinal function; but also infections or dysbiosis, cholestasis, and the removal of toxins from circulation.

Stages of Detoxification

The process of detoxification consists of three phases, although many people are only familiar with the first two of these. Phase I reactions involve the oxidation, reduction and hydrolysis of substances via enzymes from the cytochrome P450 (CYP450) family.(14) It is Phase I metabolism that converts many drugs into their active compounds, and converts some chemicals into more toxic metabolites. Phase II metabolism involves the conjugation of toxins, creating larger, inactive, water-soluble molecules. Phase II reactions include sulfation, glucuronidation, and glutathione conjugation (see Figure 1).(15)

Figure 1

Figure 1

Figure 1: In a healthy state, toxins are processed and removed from the cells and organs via enzymes and transporter proteins, and leave the body primarily via the skin, urine, and feces.

Phase III is often neglected in discussions of detoxification. However, it is critical. Phase III involves the process of transport and elimination of toxic substances through cellular membranes. The primary proteins that play a role in Phase III are multidrug resistance protein (MRP) 1, 2, 3, and 4, organic anion transport proteins (OATP), and P-glycoprotein (P-gp).(16) These proteins also regulate the movement of molecules through barrier tissues, such as the blood-brain barrier.(17)


The work horses for detoxification are 1) MRP1, the transmembrane transporter serving most cells in the body for exporting toxins to circulation, 2) OATP, the basolateral membrane transporter which moves toxin conjugates from the blood into the hepatocytes or renal tubule epithelial cells, and 3) MRP2, the apical transporter that moves toxin conjugates (and some bile salts) into the bile canaliculus or renal proximal tubule lumen. MRP3 and MRP4 are basolateral transporters that move toxin conjugates and bile salts from hepatocytes into the blood. All of the enzymes and transporters required for detoxification are present at a basal level, but many have increased expression in a coordinated fashion when stimulated by drug or toxin exposure.(18,19)

Although the process of detoxification occurs in every cell of the body, the liver, kidneys, and intestines are primary tissues in which higher levels of detoxification occur.(20) Many are familiar with the importance of the liver and kidneys as organs of detoxification, yet neglect awareness of the role of the intestines, the mucosal lining of which expresses high levels of the proteins important for all phases of detoxification.(21,22) When any of these systems are impaired, a backup in processing of toxins will occur, with a greater burden being placed on other organs.

Nrf2: The Cellular Detoxification On-Switch

Nrf2 (short for nuclear factor E2-related factor) is a cellular switch that orchestrates antioxidant, detoxification, and cellular defenses. Nrf2 is present in the cytosol of the cell (see Figure 2), and responds to oxidative stress by translocating to the nucleus and binding to the promoter region of genes that encode the transcription of critical components of detoxification known as the antioxidant response element (ARE).(23) In addition to elevated levels of reactive oxygen species (ROS), the Nrf2/ARE pathway is activated by a reduced cellular antioxidant capacity, and by exposure to toxic substances like air pollution and heavy metals.(24,25)

Figure 2

Figure 2: Oxidative stress causes Nrf2 to dissociate from binding protein (Keap1) in the cytosol and translocate to the nucleus where it binds the promoter region (ARE), leading to transcription of detoxification enzymes and proteins. Various substances have been shown to have an inhibitory or inducing effect on the Nrf2/ARE pathway.

 

When activated, the Nrf2/ARE pathway can switch on over 200 genes that help the cell generate protective molecules (see Fig 2).(26) This includes antioxidant elements, detoxification enzymes, and proteins required for glutathione synthesis and recycling such as glutathione S-transferase (GST), glutathione reductase (GSR), and glutathione peroxidase (GPX).(27-30) Nrf2 also upregulates proteins responsible for Phase III detoxification (P-gp, BCRP, and MRP2) and the transfer of toxic substances out of the cell and central nervous system.

Factors that Impede Detoxification
Nrf2/ARE Pathway. Studies have shown that the ability to upregulate Nrf2 and its antioxidant-supporting action declines with age, which may be one reason the elderly are more susceptible to damage from environmental pollutants.(31) Ochratoxin A, one of the most common mycotoxins found in foods and water-damaged houses, acts as a Nrf2 inhibitor (see Figure 2).(32) Indoxyl sulfate, a uremic toxin that is increased in the blood with chronic kidney disease and exposure to toxic heavy metals such as cadmium, also acts as a Nrf2 inhibitor, further contributing to accelerated renal damage at the level of the tubules.(33-35)

Endotoxin. Lipopolysaccharide (LPS), also known as endotoxin, is associated with the outer membrane of gram-negative bacteria. Increased intestinal permeability, aka "leaky gut," allows for increased translocation of LPS from the gut into circulation. Damage to the intestinal barrier is common and can occur with infection, surgery, stress, intense exercise, celiac disease, food allergies, non-steroidal anti-inflammatory drugs (NSAIDs), and alcohol use (see Figure 3).(36-38) Toxins, including heavy metals, pesticides, and herbicides such as glyphosate, also have been shown to lead to inflammation and/or increased permeability.(39-43) With exposure to mercury, inflammation and increased intestinal permeability may occur due to oxidative stress and glutathione depletion.(44)

Figure 3

Figure 3: Gastrointestinal inflammation and increased intestinal permeability allow for endotoxin (LPS) to be released from bacteria in the gut into circulation. Endotoxin and related inflammatory cytokines block detoxification pathways by downregulating the detoxification enzymes and Phase III transporters, as well as contributing to cholestasis and kidney damage.

Endotoxin and the associated inflammation leads to glutathione depletion, further contributing to cellular damage as toxins are no longer efficiently transported out of the cells, or protected from oxidative stress.(45,46) Exposure to endotoxin and the cascade of inflammatory cytokines it triggers also has the effect of downregulating expression of some of the important CYP enzymes and Phase III transporters.(47,48) Endotoxin exposure has a dramatic effect on the urinary elimination of mercury, acting synergistically with the heavy metal to further induce kidney damage.(49) Endotoxin also has an effect of rapidly and dramatically reducing bile flow by suppressing expression and function of hepatobiliary transporters (Figure 4b).(50)

Cholestasis. Bile plays a role in the human body not only for the emulsification and digestion of fatty substances, but also regulates many critical facets of physiology including glucose and cholesterol metabolism as well as thyroid hormone activation.(51,52) Along with bile salts, the body secretes cholesterol and phospholipids as well as toxins out of the liver and into the intestines, where they either move out of the body or are reabsorbed via enterohepatic circulation. Bile salts have an impact on the gastrointestinal flora and promote normal gastrointestinal motility.(53,54) Because of these many important functions, diminished bile flow can have a serious and broad ranging impact on health.

Figure 4a

Figure 4a

Figure 4a. Normal functioning hepatocyte. FXR remains in the cytosol until activated by bile acids. Oxidative stress causes Nrf2 to dissociate from Keap1 in the cytosol and bind ARE in the nucleus, increasing transcription of detoxification-related enzymes and proteins.



Bile acids are normally secreted from hepatocytes across the canalicular membrane via the bile salt export protein (BSEP), as well as the Phase III transporter MRP2.(55) BSEP and MRP2 are from the same superfamily of transporters known as ATP-binding cassette (ABC) transporters, which also includes the Phase III transporters MRP1, MRP3, MRP4, BCRP, and P-gp. Very importantly, BSEP and MRP2 have an interdependent expression, and under normal conditions are colocalized in the apical membrane of the hepatocytes lining the bile canaliculi (see Figure 4a).(56) The binding of bile salts to nuclear bile salt receptors, including farnesoid X receptor (FXR),(57) pregnane X receptor (PXR),(58) the vitamin D receptor (VDR),(59) and possibly the xenobiotic receptor, constitutive androstane receptor (CAR),(60) increases the expression of transporters for their efflux from the cell, and also regulates their uptake and biosynthesis (see Figure 4a).(61)

The rate limiting step in bile salt excretion is transport at the canalicular membrane, as there is a high concentration gradient to overcome in order to excrete bile salts into the bile acid pool.(62) There are many factors which can inhibit or limit bile acid production and secretion. Substances such as estrogen (in excess), certain medications (including antidepressants), endotoxin, and related inflammatory cytokines are capable of inducing cholestasis by impairing the function of the bile acid transport proteins.(63-66) Though often triggered by inflammatory responses, cholestasis also induces an inflammatory response, leading to ROS- and surfactant-induced hepatocyte damage and death due to intracellular bile sale accumulation.(67,68) This failure to move bile salts into the bile canaliculus is termed intrahepatic cholestasis.

Cholestasis is toxistasis. With cholestasis, not only is there reduced biliary excretion of bile, but, due to coregulation of BSEP and MRP2, detoxification is impaired as well. There is a reduction of transport of toxins out of the cell by the Phase III proteins, a reduction of Phase II metabolism, and decreased hepatocellular synthesis of GSH, in part due to the blocking of Nrf2 binding to ARE (see Figure 4b).(69-71) In cholestasis, a decreased expression of BSEP and the Phase III transporter MRP2 is seen at the canalicular membrane.(72) The Phase II estrogen metabolite, estradiol-17β-d-glucuronide (E217βG), triggers internalization of both BSEP and MRP2,(73) while LPS causes relocation of MRP2 to the basolateral membrane.(74) Each of these factors negatively impacts the ability of the hepatocyte to transport toxins out into the bile. OATP, which serves to transport bile acids and toxins from the blood into the hepatocyte, decreases. An additional protein in the ABC transporter family, MRP3, upregulates in cholestasis, protecting the hepatocytes from toxin-related damage and death.(75) However, rather than serving to transport toxins out into the bile canaliculi as MRP2 does, it eliminates them from the cell back into the blood and neighboring cells (see Figure 4b).(76) With the additional relocation of MRP2 to the basolateral membrane there are now two pumps moving toxins and toxin conjugates back into the blood during cholestasis. This is likely the mechanism of "detox reactions" or "Herxheimer reactions" experienced during unbalanced detoxification protocols and points to therapeutic interventions to remedy those reactions or prevent them in the first place.

Shade Decker Image 4b.jpg

Figure 4b: In cholestasis, there is reduced levels of BSEP and MRP2 at the canalicular membrane due to internalization and relocation. Binding of bile salts to FXR inhibits NTCB transport of bile acids into the cell and increases transcription of BSEP and MRP3 and 4 to lower intracellular bile acid concentration.  Nrf2 is blocked from binding ARE by c-Maf/MafG, leading to reduced Phase II inactivation of toxins as well as diminished glutathione synthesis.



The kidney reflexively adapts in attempts to support bile salt removal from the blood in cholestasis by a variety of mechanisms.(77) Passive glomerular filtration increases, while at the level of the tubules, active secretion of bile increases, and tubular reabsorption of bile acids is repressed.(78) MRP2 is one of the specific proteins that have increased expression in the kidney, protecting the organism by increasing renal bile salt and toxin elimination.(79) In severe biliary obstruction, acute renal failure may occur.(80) In the intestines, with biliary obstruction, the expression of MRP2 in the enterocytes, where it also serves to transport toxins out, is dramatically reduced.(81) The intestinal reabsorption of bile also adapts in cholestasis, as a bile acid transporter that contributes substantially to enterohepatic reabsorption in the duodenum is also downregulated.(82)

Clinical Manifestations of Cholestasis/Toxistasis

A range of conditions including biliary obstruction, pregnancy, chronic viral hepatitis, cirrhosis, primary biliary cholangitis, and primary sclerosing cholangitis may lead to cholestasis. Cholestasis related to extrahepatic or intrahepatic biliary obstruction is suggested by gastrointestinal symptoms including right upper quadrant pain or tenderness which may be prolonged, epigastric tenderness, discomfort or nausea after meals, and stool changes possibly including evidence of gross fat malabsorption (see Table 1a). Symptoms often are exaggerated with consumption of meals containing a high amount of fat. General or isolated pruritis, localized to the palms or soles of feet, is common, and often worse at night or pre-menstrually in women.(83) Depending on the cause of cholestasis, there also may be symptoms of fatigue and impaired memory and concentration.(84,85) These symptoms, however, mirror those of toxemia and point to the liver "backfire" (basolateral toxin transport dominating over canalicular transport, Figure 4b) described previously as being causal. Laboratory findings and imaging which may suggest cholestasis are found in Table 1b.

  • Table 1a: Signs and Symptoms of Cholestasis
    Right upper quadrant pain or tenderness
    Discomfort or nausea after meals
    Pruritis, often worse at night
    Epigastric tenderness
    Fat malabsorption
    Stool changes, especially pale stool
    Jaundice
    Fatigue
    Impaired memory and concentration

  • Table 1b: Labs and Imaging Suggestive of Cholestasis
    Elevated serum aminotransferases(86)
    Elevated alkaline phosphatase, particularly if marked elevation with respect to aminotransferases(87)
    Elevated gamma-glutamyl transpeptidase(88)
    Elevated bilirubin
    Elevated 5'-nucleotidase(89,90)

    Elevated serum bile acids(91)
    Biliary sludge or gallstones shown on imaging

A "Push-Catch" Strategy to Maximize Liver Detoxification Pathways

Successful and effective detoxification requires not just support for the phases of detoxification, but also the implementation of a proper directionality to progressively mobilize and eliminate toxins from the cells and tissues, and then from the body as a larger entity. As described previously, much of this is controlled at the canalicular membrane of the hepatocyte; however, it also involves the efficient binding of toxins in the upper GI tract. With a rapid delivery system, such as nanoscale lipid-based deliveries, it is possible to supplement nutraceuticals to support detoxification phases and stimulate bile flow (the "push") and then, within 30 minutes, follow with solid-phase toxin binders (the "catch"). This Push-Catch strategy creates an efficient and discreet detoxification cycle that can be done once per day or can be repeated multiple times per day for more rapid detoxification. Importantly, because of the focus on directionality, "detoxification symptoms" are minimized or eliminated. Though many combinations of compounds can be used, the following are core concepts.

Glutathione Support

Glutathione is central to multiple cell functions, such as detoxification, free-radical control, immune balance, and cell growth. Because glutathione is lost during removal and elimination of mercury and other toxins from the cell, it also can become chronically depleted in settings of toxicity.(92) Many conditions such as autoimmune disease,(93) chronic infections,(94,95) and autism(96) are also associated with lower levels of glutathione, which sheds light on why individuals who experience one of these are often extremely vulnerable to additional insults and toxicants.(97) Furthermore, liver concentrations of glutathione are five-fold higher than other cells in the body. Glutathione support is thus a cornerstone of detoxification protocols.

Typical oral supplementation of glutathione has low bioavailability, and minimally impacts intracellular levels.(98) N-acetylcysteine (NAC), which supports the production of glutathione by providing the precursor L-cysteine, also has limited ability to support intracellular glutathione levels as the conversion of L-cysteine to glutathione is often poor.(99-101) Because of this, intravenous glutathione, and alternate oral glutathione delivery forms, such as liposomes and S-acetylglutathione, are often utilized.(102,103)

Another important mechanism to consider in detoxification protocols is inducing the endogenous cellular production of glutathione and the related antioxidant-supporting and chemoprotective (detoxifying) enzymes and proteins via the Nrf2/ARE pathway (see Figure 2). Sustained activation of Nrf2 has been shown to counteract hepatic injury and bilirubin elevation associated with cholestasis.(104) Although Nrf2 is maintained at a basal level, it has a half-life of approximately 15 minutes and is constantly degraded in cells not experiencing stress.(105,106) Natural substances that have been shown to induce Nrf2 include lipoic acid (especially the R-form), selenium, diindolylmethane (DIM), sulforaphane, lycopene, milk thistle, and epigallocatechin gallate (EGCG).(107-111) Importantly, lifestyle factors also have the ability to affect Nrf2 induction. Activities such as relaxation, breathing techniques, and exercise have the effect of inducing Nrf2.(112-114)

Hepatoprotection and Biliary Support

Some of the medications which improve symptoms and biochemical markers of liver injury in settings of cholestasis have mechanisms that include supporting the detoxification pathways. Ursodeoxycholic acid (UDCA), a primary medication used in settings of cholestasis, may have a protective effect via co-regulation of Phase III transporter expression. UDCA stimulates hepatic BSEP, and also co-stimulates hepatic, intestinal, and renal MRP2.(115) UDCA, as well as S-adenosylmethionine (SAMe), prevents the cholestasis-induced blockage to Nrf2/ARE binding, increasing synthesis of detoxification-related enzymes and glutathione.(71,116) Rifampicin, a medication that is used primarily as an antibiotic, but also for pruritis associated with cholestatic liver disease,117 enhances bile acid detoxification by increasing expression of CYP3A4 (Phase 1), UDP-glucuronosyltransferases (Phase 2), and MRP2 (Phase 3).(118)

Many natural substances support detoxification by improving biliary elimination of toxins. Phosphatidylcholine, the predominant phospholipid build-ing block of cellular membranes, is a crucial constituent of bile. As phosphatidylcholine comprises over 90% of the total bile phospholipids content,(119) inadequate intake contributes to impaired biliary excretion of bile and toxins, and promotes cholesterol crystallization and gallstone formation.(120) This further promotes liver damage by obstruction of the small bile ducts. Increased intake of phosphatidylcholine has been shown to enhance biliary lipid secretion, preventing cholestasis and subsequent liver damage.(121,122) Although small amounts of choline can be synthesized from methionine or serine, it is considered an essential nutrient and must be obtained from the diet.(123) A recent study showed that only 8% of US adults meet the recommended adequate intake (AI) of choline, with vegetarians, postmenopausal women, and men at greater risk of inadequacy.(124,125)

Bitter Herbs

Well known for their generally stimulating effect on digestive system function, bitter herbs play an important role in promoting adequate biliary secretion. Digestive bitters which have hepatoprotective effects and/or support the formation and elimination of bile include gentian, dandelion, myrrh, and milk thistle. Some of these botanicals also have specific mechanisms by which they have been shown to support detoxification pathways.

Gentian (Gentiana lutea) is one of the strongest herbal bitters that is often utilized in digestive bitter formulations. Gentian has been shown to have a choleretic effect, normalizing bile volume in the setting of liver injury.(126) As a liver protective agent, gentian has been observed to increase levels of GSH, GSR, GPX, and superoxide dismutase which were otherwise reduced by alcohol or acetaminophen-induced oxidative damage.(127,128)

Dandelion (Taraxacum officinale) simultaneously stimulates the production of bile by the liver (choleretic), the flow of bile into the small intestine (cholagogue), and also has hepatoprotective effects.(129) In the setting of alcohol-induced oxidative stress, supplementation with dandelion root extract has also been observed to increase hepatic antioxidant activity, including GSH, GST, GPX, and GSR.(130)

Myrrh (Commiphora myrrha) has a complex profile of use and is perhaps most recognized for its antimicrobial effect.(131,132) Myrrh also acts as an anesthetic, anti-inflammatory, antioxidant, and cholesterol-lowering agent.(133) In Ayurvedic medicine, myrrh is used as a detoxifier and female reproductive tonifying agent, helping to move stagnant blood.(134) Myrrh, and its close relative guggul (Commiphora mukul), contain molecules known as guggulsterones that have diverse biological activities. The guggulsterones are the bioactive agents responsible for the cholesterol-lowering effect, as well as anti-inflammatory and anti-oxidative properties.(135) Guggulsterones have been shown to increase the transcription of BSEP as well as induction of the detoxification-promoting nuclear transcription factor PXR.

Milk thistle (Silybum marianum) has been vastly studied for its anti-oxidative, anti-inflammatory, and hepatoprotective effects.(136) Silymarin is the active complex extracted from the seeds of the plant, with the flavonolignan silybin, also known as silibinin, being the most biologically active moiety comprising 50% to 70% of silymarin. One of the most important mechanisms by which milk thistle supports detoxification, in addition to its antioxidant effects, is via its anti-cholestatic properties.(137) Silibinin stabilizes BSEP in its hepatocyte membrane location, preventing cholestasis caused by BSEP internalization in the presence of substances like estrogen.(138) When co-administered with estrogen, silymarin was shown to prevent the estrogen-induced decrease in bile-salt dependent bile flow.(139) Silibinin and silymarin also have been shown to stimulate the nuclear bile salt receptor FXR in a dose dependent manner, which increases expression of BSEP and MRP2, and also may have other positive metabolic effects.(140)

Toxin Binders

Completing the process of detoxification requires intestinal binders for two reasons: 1) many toxins (methylmercury, cadmium, and mycotoxins being well-known, as well as others with increased intestinal permeability) are reabsorbed after excretion into the bile, and 2) endotoxin and other dysbiotic toxins derived from the gut can be prophylactically bound with non-absorbed sorbent substances like activated carbon. As translocation of endotoxin from the gastrointestinal tract to circulation not only directly causes inflammation and oxidative damage but also has a dramatic negative effect on detoxification, it is imperative to bind and remove it from the body. Supporting reduction of intestinal permeability is also an important aspect of detoxification strategies. However, because there is no universal toxin binder that has an equal affinity for all toxins (heavy metals, molds, plastics, and more), a combination of binders that span a breadth of possible toxin chemistries is necessary (see Table 2).

Klaire2.jpg

Table 2: Toxic Substances Bound by Common Binders

  • Activated charcoal: Endotoxin, mycotoxins, pesticides and herbicides, volatile organic compounds (VOCs)

  • Bentonite clay: Mycotoxins, bisphenol A (BPA), pesticides and herbicides, some metal binding, also has antibacterial activity

  • Chitosan, a molecular mimic of Welchol: Ochratoxin, polychlorinated biphenyls (PCBs), phthalates, BPA, endotoxin, metals, also has prebiotic activity

  • Thiol-functionalized silica: Heavy metals specific binder including mercury, lead, arsenic, and cadmium

Activated charcoal is well known for its ability to adsorb a wide variety of toxic substances, and is used for this purpose in many emergency settings when poisonous substances or medication overdoses have been ingested.(141) One of the most important things about charcoal is that it is very effective at binding and removing endotoxin, a major contributor to blocked detoxification pathways.(142-144) Activated charcoal also effectively adsorbs pesticides and herbicides,(145) volatile organic compounds (VOCs) such as benzene,(146) mycotoxins,(147) and the intestinal precursor to indoxyl sulfate, a uremic toxin.(148) Charcoal has also been observed to reduce pro-inflammatory cytokine production in settings of infection.(149)

Bentonite clay is particularly good at absorbing mycotoxins, including food-borne aflatoxin; aflatoxin's precursor mycotoxin sterigmatocystin, commonly found in water-damaged buildings;(150,151) zearalenone, a mycotoxin with estrogenic effects commonly found on stored grains152; and fumonisin B1, a mycotoxin most often found on corn.(153) Bentonite clay also strongly binds bisphenol A (BPA),(154) as well as pesticides and herbicides,(155,156) and cyanotoxins, a product of harmful algal blooms that may be found in contaminated drinking water or food.(157) Bentonite clay has an affinity for some heavy metals such as lead,(158) cadmium,(159) and nickel,(160) and has been shown to reduce the cadmium-induced toxicity and pro-inflammatory response in vivo as well.(161,162) Bentonite clay also has intrinsic broad-spectrum antibacterial properties and has a healing effect on the gastrointestinal lining.(163)

Derived from shellfish, chitosan is the result of enzymatic treatment of chitin, a component of the shell. As a biomaterial with use in a variety of applications including as a vaccine adjuvant, chitosan has been observed to be safe for use in individuals with shellfish allergies.(164,165) Chitosan acts similarly to the bile acid sequestrants cholestyramine (Questran) and colesevelam (Welchol),(166) preventing the absorption of lipids by effectively binding to bile salts,(167) but most importantly where detoxification is concerned, removing the many conjugated toxins excreted in the bile. One extremely harmful and common toxin, ochratoxin, a mold toxin found in many foods as well as water-damaged buildings,(168,169) is very effectively bound and removed by chitosan.(170,171) Chitosan also binds metals including mercury(172,173) as well as polychlorinated biphenyls (PCBs), phthalates,(174) and BPA.(175) Chitosan, like charcoal, also is able to bind endotoxin.(176,177) Chitosan also has a prebiotic effect, promoting the growth of Bifidobacterium and Lactobacillus.(178) Like bentonite clay, chitosan also has been demonstrated to have an antimicrobial effect.(179)

Although chitosan and bentonite clay have an ability to bind some heavy metals, they are not the most effective tools for this purpose. Thiolated resins are substances with covalently attached thiolic metal-binding groups which very tightly bind metals including lead, mercury, cadmium, and arsenic.(180,181) The use of thiolated resins dates back to the 1970s when they were used to address methylmercury (MeHg) poisoning in Iraq, and were found to significantly reduce the half-life of MeHg from 61 to 20 days, performing even better than penicillamine, a medical metal-chelating agent.(182,183) The thiol-functionalized silica intercepts MeHg and other metals trapped in enterohepatic circulation, binding them and escorting them out of the intestines.(184)

Because binders act locally in the gastrointestinal tract, they allow tissue-bound toxins such as metals to safely drain into the blood at a natural rate. This contrasts with many blood metal chelating agents, which may increase circulatory levels of metals and place a greater burden on the kidneys and liver in the process of elimination.(185) With gastrointestinal binders, the work of the liver and kidneys to eliminate toxic substances including metals is diminished as enterohepatic reabsorption is interrupted. The ability of charcoal and chitosan to block initial absorption of endotoxin is an important aspect of the role of binders in effective detoxification protocols.

Nanoscale Delivery Systems Optimize Detoxification

In order to appropriately time the cellular and hepatobiliary flushing of toxins with a gastrointestinal binder to properly bind and eliminate them, a nutritional delivery system with rapid uptake and cellular delivery is necessary. Lipid nanoparticle delivery systems pose a feasible solution, as appropriately designed lipid-based vesicles have the potential for rapid uptake into circulation and greatly increased cellular delivery.(186) However, not all liposomal and nanoemulsifed particles are able to deliver these benefits, as only appropriately sized particles with properly designed surface chemistry enable rapid intraoral absorption and enhanced cellular delivery.

Particle size has a dramatic effect on systemic and cellular absorption, the capacity of the vesicles to extravasate from blood vessels and permeate into tissues, and the ability to evade immune system clearance.(187) The consideration of each of these factors has led to the optimal sizing of lipid nanoparticle delivery systems in the range of 50 to 100 nm. As capillary pore size ranges from only 6 to 12 nm in endocrine glands to 50 to 180 nm in the discontinuous leaky capillaries,(188) it is obvious that only the small liposomes will be able to permeate into the tissues through these openings.

Surface modifying techniques also can be used to improve the ability of lipid nanoparticles to traverse through the blood vessel endothelium and be absorbed by the tissues.(189) Particles utilizing surface hydration technology have been observed to dramatically increase intraoral absorption of liposomal particles, as shown in Figure 5, which compares delivery of B12 with and without surface modification to that of non-liposomal B12. Intelligently-designed surface modifications of lipid nanoparticles have also been shown to prolong the time a therapeutic agent is in circulation, reducing clearance by the mononuclear phagocyte system.(190)

Figure 5

Although absorption of the very small lipid nanovesicles primarily occurs intraorally, there always will be a percentage of the nanoparticle-containing liquid which is swallowed and experiences lower gastrointestinal absorption. In this setting, the lipid vesicle serves to protect the substances which it contains from degradation by the harsh gastric juices. These particles are absorbed via the lymphatics, which also allows for them to bypass first-pass hepatic metabolism, increasing bioavailability.(191)


In addition to their use in clinical applications for the delivery of drugs including anti-cancer, anti-fungal, and anti-inflammatory medications,(192,193) lipid nanoparticle delivery systems have been shown to dramatically improve absorption of a variety of natural substances such as DIM and milk thistle, which otherwise have poor bioavailability.(194-196) Liposomal delivery systems are becoming increasingly popular for delivery of substances such as glutathione because they protect it from breakdown in the digestive system, and, in cell culture studies, have been shown to dramatically increase intracellular delivery 100-fold over non-liposomal formats.(197) Because optimally-sized liposomes with intelligent surface modifications prolong the time the therapeutic core remains in circulation, they are ideal for the delivery of many substances for which a prolonged systemic effect is desirable. Phosphatidylcholine, which forms the external membrane of lipid nanoparticles, also nourishes cellular membranes by providing necessary phospholipids for cellular repair.(198)

Although many products claim improved bioavailability via liposomal delivery, few are able to truly deliver the increased absorption these systems are capable of. However, with appropriately engineered lipid nanoparticle delivery systems, the rate of absorption, cellular delivery, and bioavailability of many medications and natural substances can be dramatically enhanced.

~

Dr. Christopher Shade, PhD, founder and CEO of Quicksilver Scientific, specializes in the biological, environmental, and analytical chemistry of mercury in all its forms and their interactions with sulfur compounds, particularly glutathione and its enzyme system. He has patented analytical systems for mercury speciation (separation of different forms of mercury), founded the only clinical lab in the world offering mercury speciation in human samples, and has designed cutting edge systems of nutraceuticals for detoxification and antioxidant protection, including advanced phospholipid delivery systems for both water- and fat-soluble compounds. Dr. Shade is regularly sought out to speak as an educator on the topics of mercury, environmental toxicities, neuroinflammation, immune dysregulation, and the human detoxification system for practitioners and patients in the United States and internationally.

Dr. Carrie Decker, ND, graduated with honors from the National College of Natural Medicine (now the National University of Natural Medicine) in Portland, Oregon. Dr. Decker sees patients at her office in Portland, OR, as well as remotely, with a focus on gastrointestinal disease, mood imbalances, eating disorders, autoimmune disease, and chronic fatigue. Prior to becoming a naturopathic physician, Dr. Decker was an engineer and obtained graduate degrees in biomedical and mechanical engineering from the University of Wisconsin-Madison and University of Illinois at Urbana-Champaign respectively. Dr. Decker continues to enjoy academic research and writing, and uses these skills to support integrative medicine education as a writer and contributor to various resources.

Contact information:
Christopher Shade, PhD
christopher.shade@quicksilverscientific.com
1376 Miners Dr., Ste. 103
Lafayette, Colorado 80026

Carrie Decker, ND
BlessedThistleND@gmail.com
728 NE Dekum St.
Portland, Oregon 97211

References

 

1. Terzano C, et al. Air pollution ultrafine particles: toxicity beyond the lung. Eur Rev Med Pharmacol Sci. 2010 Oct;14(10):809-21.

2. Lelieveld J, et al. The contribution of outdoor air pollution sources to premature mortality on a global scale. Nature. 2015 Sep 17;525(7569):367-71.

3. Sorg TJ, Chen AS, Wang L. Arsenic species in drinking water wells in the USA with high arsenic concentrations. Water Res. 2014 Jan 1;48:156-69. 

4. Villaneuva CM, et al. Assessing Exposure and Health Consequences of Chemicals in Drinking Water: Current State of Knowledge and Research Needs. Environ Health Perspect. 2014 Mar; 122(3): 213–221. 

5. Thapa K, Pant BR. Pesticides in vegetable and food commodities: environment and public health concern. J Nepal Health Res Counc. 2014 Sep-Oct;12(28):208-10.

6. Verger PJ, Boobis AR. Global food supply. Reevaluate pesticides for food security and safety. Science. 2013 Aug 16;341(6147):717-8.

7. Zalko D, et al. Viable skin efficiently absorbs and metabolizes bisphenol A. Chemosphere. 2011 Jan;82(3):424-30.

8. Nguyen T, Nioi P, Pickett CB. The Nrf2-antioxidant response element signaling pathway and its activation by oxidative stress. J Biol Chem. 2009 May 15;284(20):13291-5.

9. Carocci A, et al. Mercury toxicity and neurodegenerative effects. Rev Environ Contam Toxicol. 2014;229:118.

10. Irigaray P, et al. Lifestyle-related factors and environmental agents causing cancer: an overview. Biomed Pharmacother. 2007 Dec;61(10):640-58. 

11. Bhatnagar A, et al. Environmental cardiology: studying mechanistic links between pollution and heart disease. Circ Res. 2006 Sep 29;99(7):692-705. 

12. Mendiola J, et al. Exposure to environmental toxins in males seeking infertility treatment: a case-controlled study. Reprod Biomed Online. 2008 Jun;16(6):842-50. 

13. Diaz-Sanchez D, et al. Diesel fumes and the rising prevalence of atopy: an urban legend? Curr Allergy Asthma Rep. 2003 Mar;3(2):146-52. 

14. Grant DM. Detoxification pathways in the liver. J Inherit Metab Dis. 1991;14(4):421-30. 

15. Zamek-Gliszczynski MJ, et al. Integration of hepatic drug transporters and phase II metabolizing enzymes: mechanisms of hepatic excretion of sulfate, glucuronide, and GSH metabolites. Eur J Pharm Sci. 2006 Apr;27(5):447-86. 

16. Leslie EM, Deeley RG, Cole SP. Multidrug resistance proteins: role of P-glycoprotein, MRP1, MRP2, and BCRP (ABCG2) in tissue defense. Toxicol Appl Pharmacol. 2005 May 1;204(3):216-37. 

17. Miller DS. Regulation of P-glycoprotein and other ABC drug transporters at the blood-brain barrier. Trends Pharmacol Sci. 2010 Jun;31(6):246-54.

18. Rushmore TH, Kong AN. Pharmacogenomics, regulation and signaling pathways of phase I and II drug metabolizing enzymes. Curr Drug Metab. 2002 Oct;3(5):481-90. 

19. Catania VA, et al. Co-regulation of expression of phase II metabolizing enzymes and multidrug resistanceassociated protein 2. Ann Hepatol. 2004 Jan-Mar;3(1):11-7. 

20. Hilgendorf C, et al. Expression of thirty-six drug transporter genes in human intestine, liver, kidney, and organotypic cell lines. Drug Metab Dispos. 2007 Aug;35(8):1333-40.

21. Doherty MM, Charman WN. The mucosa of the small intestine: how clinically relevant as an organ of drug metabolism? Clin Pharmacokinet. 2002;41(4):235-53. 

22. Berggren S, et al. Gene and protein expression of P-glycoprotein, MRP1, MRP2, and CYP3A4 in the small and large human intestine. Mol Pharm. 2007 Mar-Apr;4(2):252-7. 

23. Petri S, Körner S, Kiaei M. Nrf2/ARE Signaling Pathway: Key Mediator in Oxidative Stress and Potential Therapeutic Target in ALS. Neurol Res Int. 2012;2012:878030. 

24. Risom L, Møller P, Loft S. Oxidative stress-induced DNA damage by particulate air pollution. Mutat Res. 2005 Dec 30;592(1-2):119-37. 

25. Simmons SO, Fan CY, Yeoman K, et al. NRF2 Oxidative Stress Induced by Heavy Metals is Cell Type Dependent. Curr Chem Genomics. 2011;5:1-12. 

26. Bruni F, Polosa PL, Galadeta MN. Nuclear Respiratory Factor 2 Induces the Expression of Many but Not All Human Proteins Acting in Mitochondrial DNA Transcription and Replication. J Biol Chem. 2010 February 5; 285(6): 3939–3948. 

27. Nguyen T, Nioi P, Pickett CB. The Nrf2-antioxidant response element signaling pathway and its activation by oxidative stress. J Biol Chem. 2009 May 15;284(20):13291-5. 

28. Itoh K, et al. An Nrf2/small Maf heterodimer mediates the induction of phase II detoxifying enzyme genes through antioxidant response elements. Biochem Biophys Res Commun. 1997 Jul 18;236(2):313-22. 

29. Harvey CJ, Thimmulappa RK, Singh A, et al. Nrf2-regulated glutathione recycling independent of biosynthesis is critical for cell survival during oxidative stress. Free Radic Biol Med. 2009 Feb 15;46(4):443-53. 

30. Singh A, Rangasamy T, Thimmulappa RK, et al. Glutathione peroxidase 2, the major cigarette smokeinducible isoform of GPX in lungs, is regulated by Nrf2. Am J Respir Cell Mol Biol. 2006 Dec;35(6):639-50. 

31. Zhang H, et al. Nrf2-regulated phase II enzymes are induced by chronic ambient nanoparticle exposure in young mice with age-related impairments. Free Radic Biol Med. 2012 May 1;52(9):2038-46. 

32. Limonciel A, Jennings P. A review of the evidence that ochratoxin A is an Nrf2 inhibitor: implications for nephrotoxicity and renal carcinogenicity. Toxins (Basel). 2014 Jan 20;6(1):371-9. 

33. Hung SC, et al. Indoxyl Sulfate: A Novel Cardiovascular Risk Factor in Chronic Kidney Disease. J Am Heart Assoc. 2017 Feb 7;6(2). 

34. Lan Z, Bi KS, Chen XH. Ligustrazine attenuates elevated levels of indoxyl sulfate, kidney injury molecule1 and clusterin in rats exposed to cadmium. Food Chem Toxicol. 2014 Jan;63:62-8. 

35. Bolati D, et al. Indoxyl sulfate, a uremic toxin, downregulates renal expression of Nrf2 through activation of NF-κB. BMC Nephrol. 2013 Mar 4;14:56. 

36. Riddington DW, Venkatesh B, Boivin CM, et al. Intestinal permeability, gastric intramucosal pH, and systemic endotoxemia in patients undergoing cardiopulmonary bypass. JAMA. 1996 Apr 3;275(13):1007-12.

37. Lindén SK, et al. Mucin dynamics in intestinal bacterial infection. PLoS One. 2008;3(12):e3952. 

38. Groschwitz KR, Hogan SP. Intestinal barrier function: molecular regulation and disease pathogenesis. J Allergy Clin Immunol. 2009 Jul;124(1):3-20. 

39. Vasiluk L, Pinto LJ, Moore MM. Oral bioavailability of glyphosate: studies using two intestinal cell lines. Environ Toxicol Chem. 2005 Jan;24(1):153-60.

40. Anton PM, et al. Chronic ingestion of a potential food contaminant induces gastrointestinal inflammation in rats: role of nitric oxide and mast cells. Dig Dis Sci. 2000 Sep;45(9):1842-9.

41. Claus SP, Guillou H, Ellero-Simatos S. The gut microbiota: a major player in the toxicity of environmental pollutants? Biofilms & Microbiomes. 2016 May 4. 

42. Awad WA, Hess C, Hess M. Enteric Pathogens and Their Toxin-Induced Disruption of the Intestinal Barrier through Alteration of Tight Junctions in Chickens. Toxins (Basel). 2017 Feb 10;9(2).

43. Batah J, et al. Clostridium difficile flagella induce a pro-inflammatory response in intestinal epithelium of mice in cooperation with toxins. Sci Rep. 2017 Jun 12;7(1):3256

44. Vázquez M, et al. In vitro evaluation of inorganic mercury and methylmercury effects on the intestinal epithelium permeability. Food Chem Toxicol. 2014 Dec;74:349-59. 

45. Nadeem A, et al. Acute glutathione depletion leads to enhancement of airway reactivity and inflammation via p38MAPK-iNOS pathway in allergic mice. Int Immunopharmacol. 2014 Sep;22(1):222-9.

46. Carbonell LF, et al. Depletion of liver glutathione potentiates the oxidative stress and decreases nitric oxide synthesis in a rat endotoxin shock model. Crit Care Med. 2000 Jun 1;28(6):2002-6.

47. Tang W, et al. Endotoxin downregulates hepatic expression of P-glycoprotein and MRP2 in 2acetylaminofluorene-treated rats. Mol Cell Biol Res Commun. 2000 Aug;4(2):90-7. 

48. Kalitsky-Szirtes J, et al. Suppression of drug-metabolizing enzymes and efflux transporters in the intestine of endotoxin-treated rats. Drug Metab Dispos. 2004 Jan;32(1):20-7.

49. Rumbeiha WK, et al. Augmentation of mercury-induced nephrotoxicity by endotoxin in the mouse. Toxicology. 2000 Oct 26;151(1-3):103-16.

50. Kosters A, Karpen SJ. The role of inflammation in cholestasis: clinical and basic aspects. Semin Liver Dis. 2010 May;30(2):186-94.

51. Zhou H, Hylemon PB. Bile acids are nutrient signaling hormones. Steroids. 2014 Aug;86:62-8. 

52. Watanabe M, et al. Bile acids induce energy expenditure by promoting intracellular thyroid hormone activation. Nature. 2006 Jan 26;439(7075):484-9.

53. Islam KB, et al. Bile acid is a host factor that regulates the composition of the cecal microbiota in rats. Gastroenterology. 2011 Nov;141(5):1773-81. 

54. Hellström PM, Nilsson I, Svenberg T. Role of bile in regulation of gut motility. J Intern Med. 1995 Apr;237(4):395-402.

55. Trauner M, Boyer JL. Bile salt transporters: molecular characterization, function, and regulation. Physiol Rev. 2003;83:633–671.

56. Zinchuk VS, et al. Asynchronous expression and colocalization of Bsep and Mrp2 during development of rat liver. Am J Physiol Gastrointest Liver Physiol. 2002 Mar;282(3):G540-8.

57. Rizzo G, et al. Role of FXR in regulating bile acid homeostasis and relevance for human diseases. Curr Drug Targets Immune Endocr Metabol Disord. 2005 Sep;5(3):289-303.

58. Xie W, et al. An essential role for nuclear receptors SXR/PXR in detoxification of cholestatic bile acids. Proc Natl Acad Sci U S A. 2001 Mar 13;98(6):3375-80.

59. Makishima M, et al. Vitamin D receptor as an intestinal bile acid sensor. Science. 2002 May 17;296(5571):1313-6.

60. Guo GL, et al. Complementary roles of farnesoid X receptor, pregnane X receptor, and constitutive androstane receptor in protection against bile acid toxicity. J Biol Chem. 2003 Nov 14;278(46):45062-71. 

61. Boyer JL. New perspectives for the treatment of cholestasis: lessons from basic science applied clinically. J Hepatol. 2007 Mar;46(3):365-71.

62. Trauner M, Boyer JL. Bile salt transporters: molecular characterization, function, and regulation. Physiol Rev. 2003 Apr;83(2):633-71.

63. Stieger B, et al. Drug- and estrogen-induced cholestasis through inhibition of the hepatocellular bile salt export pump (Bsep) of rat liver. Gastroenterology. 2000 Feb;118(2):422-30.

64. Milkiewicz P, et al. Antidepressant induced cholestasis: hepatocellular redistribution of multidrug resistant protein (MRP2). Gut. 2003 Feb;52(2):300-3.

65. Rodríguez-Garay EA. Cholestasis: human disease and experimental animal models. Ann Hepatol. 2003 OctDec;2(4):150-8.

66. Whiting JF, et al. Tumor necrosis factor-alpha decreases hepatocyte bile salt uptake and mediates endotoxin-induced cholestasis. Hepatology. 1995 Oct;22(4 Pt 1):1273-8.

67. Copple BL, Jaeschke H, Klaassen CD. Oxidative stress and the pathogenesis of cholestasis. Semin Liver Dis. 2010 May;30(2):195-204.

68. Kosters A, Karpen SJ. The role of inflammation in cholestasis: clinical and basic aspects. Semin Liver Dis. 2010 May;30(2):186-94.

69. Yang H, et al. Dysregulation of glutathione synthesis during cholestasis in mice: molecular mechanisms and therapeutic implications. Hepatology. 2009 Jun;49(6):1982-91.

70. Dietrich CG, et al. Influence of biliary cirrhosis on the detoxification and elimination of a food derived carcinogen. Gut. 2004 Dec;53(12):1850-5.

71. Yang H, et al. Dysregulation of glutathione synthesis during cholestasis in mice: molecular mechanisms and therapeutic implications. Hepatology. 2009 Jun;49(6):1982-91.

72. Crocenzi FA, et al. Localization status of hepatocellular transporters in cholestasis. Front Biosci (Landmark Ed). 2012 Jan 1;17:1201-18.

73. Crocenzi FA, et al. Estradiol-17beta-D-glucuronide induces endocytic internalization of Bsep in rats. Am J Physiol Gastrointest Liver Physiol. 2003 Aug;285(2):G449-59.

74. Zinchuk V, Zinchuk O, Okada T. Experimental LPS-induced cholestasis alters subcellular distribution and affects colocalization of Mrp2 and Bsep proteins: a quantitative colocalization study. Microsc Res Tech. 2005 Jun 1;67(2):65-70.

75. Zelcer N, et al. Transport of bile acids in multidrug-resistance-protein 3-overexpressing cells co-transfected with the ileal Na+-dependent bile-acid transporter. Biochem J. 2003 Jan 1;369(Pt 1):23-30.

76. Donner MG, Keppler D. Up-regulation of basolateral multidrug resistance protein 3 (Mrp3) in cholestatic rat liver. Hepatology. 2001 Aug;34(2):351-9.

77. Brandoni A, et al.  Expression and function of renal and hepatic organic anion transporters in extrahepatic cholestasis. World J Gastroenterol. 2012 Nov 28;18(44):6387-97.

78. Zollner G, et al. Role of nuclear receptors in the adaptive response to bile acids and cholestasis: pathogenetic and therapeutic considerations. Mol Pharm. 2006 May-Jun;3(3):231-51.

79. Lee J, et al. Adaptive regulation of bile salt transporters in kidney and liver in obstructive cholestasis in the rat. Gastroenterology. 2001 Dec;121(6):1473-84. 

80. Kramer HJ. Impaired renal function in obstructive jaundice: roles of the thromboxane and endothelin systems. Nephron. 1997;77(1):1-12. 

81. Dietrich CG, et al. Consequences of bile duct obstruction on intestinal expression and function of multidrug resistance-associated protein 2. Gastroenterology. 2004 Apr;126(4):1044-53.

82. Hruz P, et al. Adaptive regulation of the ileal apical sodium dependent bile acid transporter (ASBT) in patients with obstructive cholestasis. Gut. 2006 Mar;55(3):395-402.

83. Bunchorntavakul C, Reddy KR. Pruritus in chronic cholestatic liver disease. Clin Liver Dis. 2012 May;16(2):331-46. 

84. Newton JL, et al. Cognitive impairment in primary biliary cirrhosis: symptom impact and potential etiology. Hepatology. 2008 Aug;48(2):541-9.

85. Elliott C, et al. Functional impairment in alcoholic liver disease and non-alcoholic fatty liver disease is significant and persists over 3 years of follow-up. Dig Dis Sci. 2013 Aug;58(8):2383-91.

86. Kim YJ. [Interpretation of liver function tests]. Korean J Gastroenterol. 2008 Apr;51(4):219-24.

87. Hatoff DE, Hardison WG. Induced synthesis of alkaline phosphatase by bile acids in rat liver cell culture. Gastroenterology. 1979 Nov;77(5):1062-7. 

88. Ellis G, Worthy E, Goldberg DM. Lack of value of serum gamma-glutamyl transferase in the diagnosis of hepatobiliary disease. Clin Biochem. 1979 Aug;12(4):142-5.

89. Hill PG, Sammons HG. An assessment of 5'-nucleotidase as a liver-function test. Q J Med. 1967 Oct;36(144):457-68.

90. Righetti A, Kaplan MM. Disparate responses of serum and hepatic alkaline phosphatase and 5' nucleotidase to bile duct obstruction in the rat. Gastroenterology. 1972 May;62(5):1034-9.

91. Heikkinen J, et al. Changes in serum bile acid concentrations during normal pregnancy, in patients with intrahepatic cholestasis of pregnancy and in pregnant women with itching. Br J Obstet Gynaecol. 1981 Mar;88(3):240-5.

92. Patrick L. Mercury toxicity and antioxidants: Part 1: role of glutathione and alpha-lipoic acid in the treatment of mercury toxicity. Altern Med Rev. 2002 Dec;7(6):456-71. 

93. Perricone C, et al. Glutathione: a key player in autoimmunity. Autoimmun Rev. 2009 Jul;8(8):697-701. 

94. de Quay B, Malinverni R, Lauterburg BH. Glutathione depletion in HIV-infected patients: role of cysteine deficiency and effect of oral N-acetylcysteine. AIDS. 1992 Aug;6(8):815-9.

95. Westerveld GJ, et al. Antioxidant levels in the nasal mucosa of patients with chronic sinusitis and healthy controls. Arch Otolaryngol Head Neck Surg. 1997 Feb;123(2):201-4.

96. Omata Y, et al. Decreased zinc availability affects glutathione metabolism in neuronal cells and in the developing brain. Toxicol Sci. 2013 May;133(1):90-100.

97. Exner R, et al. Therapeutic potential of glutathione. Wiener Klinische Wochenschrift. 2000 Jul;112(14):6106. 

98. Witschi A, et al. The systemic availability of oral glutathione. Eur J Clin Pharmacol. 1992;43(6):667-9.

99. Rushworth GF, Megson IL. Existing and potential therapeutic uses for N-acetylcysteine: the need for conversion to intracellular glutathione for antioxidant benefits. Pharmacol Ther. 2014 Feb;141(2):150-9. 

100. Lu, S.C. Regulation of hepatic glutathione synthesis: current concepts and controversies. FASEB J. 1999, 13, 1169–1183

101. Gibson KR, et al. Evaluation of the antioxidant properties of N-acetylcysteine in human platelets: prerequisite for bioconversion to glutathione for antioxidant and antiplatelet activity. J Cardiovasc Pharmacol. 2009 Oct;54(4):319-26. 

102.  Vogel JU, et al. Effects of S-acetylglutathione in cell and animal model of herpes simplex virus type 1 infection. Med Microbiol Immunol. 2005 Jan;194(1-2):55-9. 

103.  Zeevalk GD, Bernard LP, Guilford FT. Liposomal-glutathione provides maintenance of intracellular glutathione and neuroprotection in mesencephalic neuronal cells. Neurochem Res. 2010 Oct;35(10):1575-87.

104. Okada K, et al. Nrf2 counteracts cholestatic liver injury via stimulation of hepatic defense systems. Biochem Biophys Res Commun. 2009 Nov 20;389(3):431-6.

105. Stewart D, et al. Degradation of transcription factor Nrf2 via the ubiquitin-proteasome pathway and stabilization by cadmium. J Biol Chem. 2003 Jan 24;278(4):2396-402. 

106. Itoh K, Tong KI, Yamamoto M. Molecular mechanism activating Nrf2-Keap1 pathway in regulation of adaptive response to electrophiles. Free Radic Biol Med. 2004 May 15;36(10):1208-13. 

107. Suh JH, et al. Decline in transcriptional activity of Nrf2 causes age-related loss of glutathione synthesis, which is reversible with lipoic acid. Proc Natl Acad Sci U S A. 2004 Mar 9;101(10):3381-6.

Researched Nutritionals 2.jpg

108. Zhang C, et al. Selenium triggers Nrf2-mediated protection against cadmium-induced chicken hepatocyte autophagy and apoptosis. Toxicol In Vitro. 2017 Oct;44:349-356. 

109. Zhao F, et al. Silymarin attenuates paraquat-induced lung injury via Nrf2-mediated pathway in vivo and in vitro. Clin Exp Pharmacol Physiol. 2015 Jul 14.

110. Surh YJ, Kundu JK, Na HK. Nrf2 as a master redox switch in turning on the cellular signalling involved in the induction of cytoprotective genes by some chemopreventive phytochemicals. Planta Med. 2008 Oct;74(13):1526-39. 

111. Saw CL, et al. Pharmacodynamics of dietary phytochemical indoles I3C and DIM: Induction of Nrf2mediated phase II drug metabolizing and antioxidant genes and synergism with isothiocyanates. Biopharm Drug Dispos. 2011 Jul;32(5):289-300. 

112. Laudenslager ML, et al. A randomized controlled pilot study of inflammatory gene expression in response to a stress management intervention for stem cell transplant caregivers. J Behav Med. 2016 Apr;39(2):346-54. 

113. Tsou YH, et al. Treadmill exercise activates Nrf2 antioxidant system to protect the nigrostriatal dopaminergic neurons from MPP+ toxicity. Exp Neurol. 2015 Jan;263:50-62. 

114. Li T, et al. Effects of different exercise durations on Keap1-Nrf2-ARE pathway activation in mouse skeletal muscle. Free Radic Res. 2015 Oct;49(10):1269-74. 

115. Zollner G, et al. Role of nuclear bile acid receptor, FXR, in adaptive ABC transporter regulation by cholic and ursodeoxycholic acid in mouse liver, kidney and intestine. J Hepatol. 2003 Oct;39(4):480-8.

116. Yang H, et al. Induction of avian musculoaponeurotic fibrosarcoma proteins by toxic bile acid inhibits expression of glutathione synthetic enzymes and contributes to cholestatic liver injury in mice. Hepatology. 2010 Apr;51(4):1291-301.

117. Prince MI, Burt AD, Jones DE. Hepatitis and liver dysfunction with rifampicin therapy for pruritus in primary biliary cirrhosis. Gut. 2002 Mar;50(3):436-9.

118. Marschall HU, et al. Complementary stimulation of hepatobiliary transport and detoxification systems by rifampicin and ursodeoxycholic acid in humans. Gastroenterology. 2005 Aug;129(2):476-85.

119. Hişmioğullari AA, Bozdayi AM, Rahman K. Biliary lipid secretion. Turk J Gastroenterol. 2007 Jun;18(2):65-70.

120. Morita SY, Terada T. Molecular mechanisms for biliary phospholipid and drug efflux mediated by ABCB4 and bile salts. Biomed Res Int. 2014;2014:954781.  

121. Chanussot F, Benkoël L. Prevention by dietary (n-6) polyunsaturated phosphatidylcholines of intrahepatic cholestasis induced by cyclosporine A in animals. Life Sci. 2003 Jun 13;73(4):381-92.

122. Karaman A, et al. Protective effect of polyunsaturated phosphatidylcholine on liver damage induced by biliary obstruction in rats. J Pediatr Surg. 2003 Sep;38(9):1341-7.

123. Canty DJ, Zeisel SH. Lecithin and choline in human health and disease. Nutr Rev. 1994 Oct;52(10):32739.

124. Wallace TC, Fulgoni VL. Usual Choline Intakes Are Associated with Egg and Protein Food Consumption in the United States. Nutrients. 2017 Aug 5;9(8).

125. Zeisel SH. Gene response elements, genetic polymorphisms and epigenetics influence the human dietary requirement for choline. IUBMB Life. 2007 Jun;59(6):380-7.

126. Oztürk N, et al. Choleretic activity of Gentiana lutea ssp. symphyandra in rats. Phytomedicine. 1998 Aug;5(4):283-8.

127. Lian LH, et al. Gentiana manshurica Kitagawa reverses acute alcohol-induced liver steatosis through blocking sterol regulatory element-binding protein-1 maturation. J Agric Food Chem. 2010 Dec 22;58(24):13013-9

128. Wang AY, et al. Gentiana manshurica Kitagawa prevents acetaminophen-induced acute hepatic injury in mice via inhibiting JNK/ERK MAPK pathway. World J Gastroenterol. 2010 Jan 21;16(3):384-91.

129. Schütz K, Carle R, Schieber A. Taraxacum--a review on its phytochemical and pharmacological profile. J Ethnopharmacol. 2006 Oct 11;107(3):313-23.

130. You Y, et al. In vitro and in vivo hepatoprotective effects of the aqueous extract from Taraxacum officinale (dandelion) root against alcohol-induced oxidative stress. Food Chem Toxicol. 2010 Jun;48(6):16327.

131. Dolara P, et al. Local anaesthetic, antibacterial and antifungal properties of sesquiterpenes from myrrh. Planta Med. 2000 May;66(4):356-8.

132. Sheir Z, et al. A safe, effective, herbal antischistosomal therapy derived from myrrh. Am J Trop Med Hyg. 2001 Dec;65(6):700-4.

133. Shen T, Li GH, Wang XN, Lou HX. The genus Commiphora: a review of its traditional uses, phytochemistry and pharmacology. J Ethnopharmacol. 2012 Jul 13;142(2):319-30.

134. Frawley D, Lad V. The Yoga of Herbs: An Ayurvedic Guide to Herbal Medicine. Lotus Press, 1986.

135. Shah R, Gulati V, Palombo EA. Pharmacological properties of guggulsterones, the major active components of gum guggul. Phytotherapy Research. 2012 Nov;26(11):1594-605.

136. Abenavoli L, et al. Milk thistle in liver diseases: past, present, future. Phytother Res. 2010 Oct;24(10):1423-32.

137. Crocenzi FA, Roma MG. Silymarin as a new hepatoprotective agent in experimental cholestasis: new possibilities for an ancient medication. Curr Med Chem. 2006;13(9):1055-74.

138. Crocenzi FA, et al. Silibinin prevents cholestasis-associated retrieval of the bile salt export pump, Bsep, in isolated rat hepatocyte couplets: possible involvement of cAMP. Biochem Pharmacol. 2005 Apr 1;69(7):111320.

Klaire1017.jpg

139. Crocenzi FA, et al. Beneficial effects of silymarin on estrogen-induced cholestasis in the rat: a study in vivo and in isolated hepatocyte couplets. Hepatology. 2001 Aug;34(2):329-39.

140. Gu M, et al. Silymarin Ameliorates Metabolic Dysfunction Associated with Diet-Induced Obesity via Activation of Farnesyl X Receptor. Front Pharmacol. 2016 Sep 28;7:345.

141. Position statement and practice guidelines on the use of multi-dose activated charcoal in the treatment of acute poisoning. American Academy of Clinical Toxicology; European Association of Poisons Centres and Clinical Toxicologists. J Toxicol Clin Toxicol. 1999;37(6):731-51.

142. Nolan JP, et al. Endotoxin binding by charged and uncharged resins. Proc Soc Exp Biol Med. 1975 Jul;149(3):766-70.

143. Du XN, et al. Effect of activated charcoal on endotoxin adsorption. Part I. An in vitro study. Biomater Artif Cells Artif Organs. 1987;15(1):229-35.

144. Pegues AS, et al. The removal of 14C labeled endotoxin by activated charcoal. Int J Artif Organs. 1979 May;2(3):153-8.

145. Zhelezova A, Cederlund H, Stenström J. Effect of Biochar Amendment and Ageing on Adsorption and Degradation of Two Herbicides. Water Air Soil Pollut. 2017;228(6):216.

146. Chiang YC, Chiang PC, Huang CP. Effects of pore structure and temperature on VOC adsorption on activated carbon. Carbon. 2001 Apr 30;39(4):523-34.

147. Monge Mdel P, et al. Activated carbons as potentially useful non-nutritive additives to prevent the effect of fumonisin B1 on sodium bentonite activity against chronic aflatoxicosis. Food Addit Contam Part A Chem Anal Control Expo Risk Assess. 2016 Jun;33(6):1043-5.

148. Schulman G. A nexus of progression of chronic kidney disease: tryptophan, profibrotic cytokines, and charcoal. J Ren Nutr. 2012 Jan;22(1):107-13.

149. de Souza JB, et al. Oral activated charcoal prevents experimental cerebral malaria in mice and in a randomized controlled clinical trial in man did not interfere with the pharmacokinetics of parenteral artesunate. PLoS One. 2010 Apr 15;5(4):e9867.

150. Tuomi T, et al. Mycotoxins in crude building materials from water-damaged buildings. Appl Enviro Microbiology. 2000 May 1;66(5):1899-904.

151. Abdel-Wahhab MA, et al. Adsorption of sterigmatocystin by montmorillonite and inhibition of its genotoxicity in the Nile tilapia fish (Oreachromis nilaticus). Mutat Res. 2005 Apr 4;582(1-2):20-7.

152. Abbès S, et al. Preventive role of phyllosilicate clay on the Immunological and Biochemical toxicity of zearalenone in Balb/c mice. Int Immunopharmacol. 2006 Aug;6(8):1251-8.

153. Mitchell NJ, et al. Calcium montmorillonite clay reduces AFB1 and FB1 biomarkers in rats exposed to single and co-exposures of aflatoxin and fumonisin. J Appl Toxicol. 2014 Jul;34(7):795-804. 

154. Park Y, et al. Bisphenol A sorption by organo-montmorillonite: implications for the removal of organic contaminants from water. Chemosphere. 2014 Jul;107:249-56.

155. Lagaly G. Pesticide–clay interactions and formulations. App Clay Sci. 2001 May 31;18(5):205-9.

156. Park Y, et al. Removal of herbicides from aqueous solutions by modified forms of montmorillonite. J Colloid Interface Sci. 2014 Feb 1;415:127-32.

157. Sukenik A, et al. Removal of cyanobacteria and cyanotoxins from lake water by composites of bentonite with micelles of the cation octadecyltrimethyl ammonium (ODTMA). Water Res. 2017 Sep 1;120:165-173

158. Naseem R, Tahir SS. Removal of Pb (II) from aqueous/acidic solutions by using bentonite as an adsorbent. Water Res. 2001 Nov 30;35(16):3982-6.

159. Pradas EG, et al. Adsorption of cadmium and zinc from aqueous solution on natural and activated bentonite. J Chemical Tech Biotech. 1994 Mar 1;59(3):289-95.

160. Vieira MG, et al. Sorption kinetics and equilibrium for the removal of nickel ions from aqueous phase on calcined Bofe bentonite clay. J Haz Mat. 2010 May 15;177(1):362-71.

161. Abbès S, et al. Inactivation of cadmium induced immunotoxicological alterations in rats by Tunisian montmorillonite clay. Int Immunopharmacol. 2007 Jun;7(6):750-60.

162. Mahrous KF, et al. Inhibition of cadmium- induced genotoxicity and histopathological changes in Nile tilapia fish by Egyptian and Tunisian montmorillonite clay. Ecotoxicol Environ Saf. 2015 Sep;119:140-7.

163. Haydel SE, Remeni CM, Willias LB. Broad-spectrum in vitro antibacterial activities of clay minerals againstantibiotic-susceptible and antibiotic-resistant bacterial pathogens. J Antimicrob Chemother 2008 Feb; 61(2): 353–361.

164. Waibel KH, et al. Safety of chitosan bandages in shellfish allergic patients. Mil Med. 2011 Oct;176(10):1153-6.

165. Baldrick P. The safety of chitosan as a pharmaceutical excipient. Regul Toxicol Pharmacol. 2010;56:290– 9.

166. Vahouny GV, et al. Comparative effects of chitosan and cholestyramine on lymphatic absorption of lipids in the rat. Am J Clin Nutr. 1983 Aug;38(2):278-84.

167. Gallaher CM, et al. Cholesterol reduction by glucomannan and chitosan is mediated by changes in cholesterol absorption and bile acid and fat excretion in rats. J Nutr. 2000 Nov;130(11):2753-9.

168. Al-Anati L, Petzinger E. Immunotoxic activity of ochratoxin A. J Vet Pharmacol Ther. 2006 Apr;29(2):7990.

169. Polizzi V, et al. JEM Spotlight: Fungi, mycotoxins and microbial volatile organic compounds in mouldy interiors from water-damaged buildings. J Enviro Monitor. 2009;11(10):1849-58.

Researched NUTRITIONALSCoreBioticAD_0118.jpg

170. Bornet A, Teissedre PL. Chitosan, chitin-glucan and chitin effects on minerals (iron, lead, cadmium) and organic (ochratoxin A) contaminants in wines. Euro Food Res Tech. 2008 Feb 1;226(4):681-9.

171. Quintela S, et al. Ochratoxin A removal from red wine by several oenological fining agents: bentonite, egg albumin, allergen-free adsorbents, chitin and chitosan. Food Addit Contam Part A Chem Anal Control Expo Risk Assess. 2012;29(7):1168-74.

172. Gerente C, et al. Application of chitosan for the removal of metals from wastewaters by adsorption— mechanisms and models review. Crit Rev Enviro Sci Tech. 2007 Jan 1;37(1):41-127. 

173. Shafaei A, Ashtiani FZ, Kaghazchi T. Equilibrium studies of the sorption of Hg (II) ions onto chitosan. Chem Eng J. 2007 Sep 15;133(1):311-6.

174. Salim CJ, Liu H, Kennedy JF. Comparative study of the adsorption on chitosan beads of phthalate esters and their degradation products. Carbo Polymers. 2010 Jul 7;81(3):640-4.

175. Dehghani MH, et al. Adsorptive removal of endocrine disrupting bisphenol A from aqueous solution using chitosan. J Enviro Chem Eng 2016 Sep 30;4(3):2647-55.

176. Davydova VN, et al. Interaction of bacterial endotoxins with chitosan. Effect of endotoxin structure, chitosan molecular mass, and ionic strength of the solution on the formation of the complex. Biochemistry (Mosc). 2000 Sep;65(9):1082-90.

177. Solov'eva T, et al. Marine compounds with therapeutic potential in gram-negative sepsis. Mar Drugs. 2013 Jun 19;11(6):2216-29.

178. Lee HW, et al. Chitosan oligosaccharides, dp 2-8, have prebiotic effect on the Bifidobacterium bifidium and Lactobacillus sp. Anaerobe. 2002 Dec;8(6):319-24.

179. Kong M, et al. Antimicrobial properties of chitosan and mode of action: a state of the art review. Int J Food Microbiol. 2010 Nov 15;144(1):51-63. 

180. Saha B, et al. Sorption of trace heavy metals by thiol containing chelating resins. Solv Extract Ion Exch. 2000 Jan 1;18(1):133-67.

181. Sangvanich T, et al. Novel oral detoxification of mercury, cadmium, and lead with thiol-modified nanoporous silica. ACS Appl Mater Interfaces. 2014 Apr 23;6(8):5483-93.

182. Clarkson TW, et al. Tests of efficacy of antidotes for removal of methylmercury in human poisoning during the Iraq outbreak. J Pharmacol Exp Ther. 1981 Jul;218(1):74-83. 

183. Rafati-Rahimzadeh M, et al. Current approaches of the management of mercury poisoning: need of the hour. Daru. 2014 Jun 2;22:46.

184. Clarkson TW, Small H, Norseth T. Excretion and Absorption of Methyl Mercury After Polythiol Resin Treatment. Arch Enviro Health: Int J. 1973 Apr 1;26(4):173-6.

185. Smith SW. The role of chelation in the treatment of other metal poisonings. J Med Toxicol. 2013 Dec;9(4):355-69.

186. Shade C. Liposomes as advanced delivery systems for nutraceuticals. Integr Med. 2016;15(1):33-36.

187. Kraft JC, et al. Emerging research and clinical development trends of liposome and lipid nanoparticle drug delivery systems. J Pharm Sci. 2014 Jan;103(1):29-52.

188. Sarin H. Physiologic upper limits of pore size of different blood capillary types and another perspective on the dual pore theory of microvascular permeability. J Angiogenes Res. 2010; 2:14.

189. Muthu MS, et al. Vitamin E TPGS coated liposomes enhanced cellular uptake and cytotoxicity of docetaxel in brain cancer cells. Int J Pharm. 2011 Dec 15;421(2):332-40.

190. Allen TM, Cullis PR. Liposomal drug delivery systems: from concept to clinical applications. Adv Drug Deliv Rev. 2013 Jan;65(1):36-48.

191. Ahn H, Park JH. Liposomal delivery systems for intestinal lymphatic drug transport. Biomater Res. 2016 Nov 23;20:36. 

192. Laouini A, et al. Preparation, characterization, and applications of liposomes: State of the art. J Colloid Sci Biotechnol. 2012;1:147–168.

193. Bulbake U, et al. Liposomal Formulations in Clinical Use: An Updated Review. Pharmaceutics. 2017 Mar 27;9(2). pii: E12.

194. Yang Z, et al. Effect of liposomes on the absorption of water-soluble active pharmaceutical ingredients via oral administration. Curr Pharm Des. 2013;19(37):6647-54. 

195. Paltsev M, et al. Comparative preclinical pharmacokinetics study of 3,3'-diindolylmethane formulations: is personalized treatment and targeted chemoprevention in the horizon? EPMA J. 2013 Dec 10;4(1):25. 

196. Yang KY, et al. Silymarin-loaded solid nanoparticles provide excellent hepatic protection: physicochemical characterization and in vivo evaluation. Int J Nanomedicine. 2013;8:3333-43. 

197. Zeevalk GD, Bernard LP, Guilford FT. Liposomal-glutathione provides maintenance of intracellular glutathione and neuroprotection in mesencephalic neuronal cells. Neurochem Res. 2010 Oct;35(10):1575-87. 

198. Spector AA, Yorek MA. Membrane lipid composition and cellular function. J Lipid Res. 1985 Sep;26(9):1015-35.