Probiotics: Supplements, Food, Pharmaceutical Industry

Swathi Putta⁎; Nagendra S. Yarla†; Dhananjaya B. Lakkappa‡; Sarat B. Imandi§; Rama Rao Malla†; Amajala K. Chaitanya†; Brahma P.V. Chari†; Silas Saka§; Rama Rao Vechalapu§; Mohammad A. Kamal,‖,#; Vadim V. Tarasov⁎⁎; Vladimir N. Chubarev⁎⁎; Korada Siva Kumar††; Gjumrakch Aliev‡‡,§§,¶¶    ⁎ University College of Pharmaceutical Sciences, Andhra University, Visakhapatnam, India
† GITAM University, Visakhapatnam, India
‡ Jain University, Ramanagara, India
§ Krishna University, Machilipatnam, India
¶ King Abdulaziz University, Jeddah, Saudi Arabia
‖ Enzymoics, Hebersham, NSW, Australia
# Novel Global Community Educational Foundation, Hebersham, NSW, Australia
⁎⁎ I.M. Sechenov First Moscow State Medical University, Moscow, Russia
†† National Institute of Food Technology Entrepreneurship and Management (NIFTEM), Haryana, India
‡‡ “GALLY” International Biomedical Research Consulting LLC, San Antonio, TX, USA
§§ University of Atlanta, Johns Creek, GA, USA
¶¶ Russian Academy of Sciences, Chernogolovka, Russia

Abstract

Advances in microbiology and molecular biology, along with the adoption of organisms not previously used as probiotics, have contributed to a growing concern about the potential safety of these microorganisms. There was an increased demand day by day for the production and improvement in the safety and quality of probiotics. Probiotics with Lactobacillus, Bifidobacterium, Streptococcus and Enterococcus are widely used as food and drugs. Different varieties of probiotic formulations are available as capsules, liquids, and powders. The strain selection and differentiation was based on the genetic complement with consumers and depends on interactions with co-administered agents or drugs. The first commercially sold dairy based probiotic was Yakult, a fermented milk with added Lactobacillus. Many more probiotic foods have come on the market, including kefir, yogurt, kombucha, kimchi, sauerkraut, other fermented foods and beverages, soft cheeses, probiotic filled drinks, sour dough breads, and milk with probiotics. Physicians have recommended probiotics as supplements to improve the efficacy of the conventional medications in such conditions as alcoholic liver diseases, urinary, respiratory and digestive tract infections, diabetes and cancer. They reduce flatulence and bloating, protect DNA, protect proteins and lipids from oxidative damage, and the maintain individual intestinal microbiota. Research is being done on the combined formulation of selected drugs with probiotics using nanotechnology to improve the patience compliance. Scientifically established health claims are a high priority to companies who seek to promote the health benefits of their particular probiotics.

Keywords

Probiotics; Functional foods; Health benefits

1 Introduction

1.1 Prebiotics

Prebiotics are the substances which reach the colon in the intact form. They are given to the animals for beneficial microbial growth in the intestine and to assist normal digestion processes. Prebiotics are short-length carbohydrates, such as fructooligosaccharides, glucooligosaccharides and inulin, which resist digestion in the upper gastrointestinal tract and are fermented in the colon to produce short-chain fatty acids, such as acetate, butyrate and propionate, and have positive effects on colonic cell growth and stability.

1.2 Probiotics

Probiotics are the live microorganisms which, when administered in adequate amounts, confer a health benefit on the host. A live microbial adjunct which has a beneficial effect on the host by modifying the host-associated or ambient microbial community, by insuring improved use of feed or by enhancing its nutrition, by enhancing the hosťs response to disease, or by improving the quality of the ambient environment.

1.3 Synbiotics

Synergistic combinations of both pro and prebiotics are called synbiotics because this approach includes a food or food supplement having both live cells of the beneficial bacteria.

2 Types of Probiotics

The microbes used as probiotics represent different types such as bacteria, yeast or mold. However, there are more common species of each such as:

2.1 Bacteria

2.2 Yeast and Molds

Saccharomyces cerevisiae, Saccharomyces bourlardii, Aspergillus niger, Aspergillus oryzae, Candida pintolopesii, Sacaromyces boulardii.

Ideal probiotic strains for this kind of application should be resistant to bile, hydrochloric acid, and pancreatic juice; be able to tolerate stomach and duodenum conditions and gastric transport; and have the ability to stimulate the immune system, thereby improving intestinal function via adhering to and colonizing the intestinal epithelium. In addition, probiotic strains must be able to survive during manufacture and storage in order to exert significant and worthwhile healthful outcomes (Lin et al., 2006). Fig. 1 demonstrates about the minimum qualification of a microbe in order to substantiate it as a probiotic (FAO/WHO, 2002). Whereas Table 1 demonstrates about the overview of various microbial species recognized as probiotics.

Fig. 1
Fig. 1 Substantiation of the word “PROBIOTIC”.

Table 1

Description of Probiotic Microorganisms
SpeciesNatureMechanismReference
Lactobacillus speciesGram-positive, facultative anaerobic or microaerophilic, rod-shapedProduction of enzymes which digest and metabolize proteins or carbohydrates, synthesize vitamin B complex, vitamin K and facilitate breakdown of bile saltsPatil and Reddy (2006)
Bifidobacterium speciesGram-Positive, non-motile, often branched anaerobic bacteria, rods shapedMetabolize lactose, generate lactic ions from lactic acid and synthesize vitamins. They also ferment indigestible carbohydrates and produce beneficial short chain fatty acidsGaldeano et al. (2007)
Streptococcus thermophilusGram-positive bacterium, and a fermentative facultative anaerobe, chain of spheresMetabolize lactose, improve lactose intolerance and antimicrobial activitySoccol et al. (2010)
Saccharomyces boulardiiGram negative, can grow under aerobic or anaerobic conditions
The diploid form is ellipsoid-shaped, haploid form is spherical shaped		Secrete proteases and other substances that breakdown bacterial enterotoxins and inhibit their binding to intestinal receptors. They also enhance vitamin production and reduce serum cholesterol levelKedar et al. (2010)

Table 1

The selection of the probiotic must be strain specific, viable and dose specific, must remain stable and viable for long period of time, have the capacity to survive through the stomach and into the intestine, and thrive in the intestine. The level of probiotics in the food that serve as delivery systems needs to be high, suggesting a minimum level of live probiotic cells should be at least 106–107 CFU/mL or CFU/g before consumption (Nualkaekul et al., 2012).

3 Functions of Gut Microbiota

4 Mechanism of Action of Probiotics

Main mechanisms of action of probiotics (Narwal and Shashi, 2011; Rastogi et al., 2011; Tiwari et al., 2012; Calafiore et al., 2012; Kumar, 2013) are the following:

5 Safety and Risk Assessment

For safety assessment of microbial species used in food and feed productions, to set priorities for the need of risk assessment. The assessment is made for a selected group of microorganisms, which if favorable, leads to the “Qualified Presumption of Safety” status (EFSA). The safety of the potential probiotic should be assessed by the minimum required tests:

6 Probiotic Therapy

6.1 Diabetes

The “gut connection” to diabetes received more attention in recent years because there is an intricate relationship between intestinal microbiota and development of metabolic diseases primarily diabetes.

Lower counts of Bifidobacterium and Faecalibacterium prausnitzii were found in diabetic individuals and both of them Gram-positive (Furet et al., 2010). Increased counts of Bacteroides ovatus and decreased Bacteroides fragilis were found in type I diabetics (Goffau et al., 2013). Type I diabetic children showed higher counts of Clostridium, Bacteroides and Veillonella, followed by lower counts of Bifidobacterium and Lactobacillus than in healthy children (Murri et al., 2013).

  • ➢ L. casei significantly decreased plasma glucose levels and inhibited the production of β-cells specific CD4 T-cells and cytokines that are the binding factors for induction of autoimmune diabetes (Matsuzaki et al., 1997).
  • ➢ L. acidophilus CCFM6, L. plantarum CCFM47, CCFM232, and L. rhamnosus GG (LGG) have the ability to affect rat intestinal α-glucosidase inhibition, short-chain fatty acids production, and utilization of prebiotics, as well as gastrointestinal tract tolerance.
  • ➢ L. acidophilus NCDC14 and L. casei NCDC19 has apparently reduced lipid peroxidation, HbA1c and ameliorated intestinal transit in diabetic rats; however, without concomitant blood glucose reduction (Yadav et al., 2008). It has also been shown that L. casei decreased the oxidative stress and suppressed the effector functions of CD4 + T cells, accompanied by reducing the proinflammatory molecules (Villarini et al., 2008).
  • ➢ L plantarum DSM15313 and L gasseri BNR17 is suggested to reduce glycaemia, improve glucose tolerance, and reduce insulin resistance (Andersson et al., 2010; Yun et al., 2009).
  • ➢ L reuteri GMNL-263 has been shown to reduce glycaemia and HbA1c levels, and to prevent renal fibrosis (Lu et al., 2010).
  • ➢ L. acidophilus La5 and B. lactis Bb12 noticeably reduce blood glucose levels, glycated hemoglobin and remarkably improve antioxidant status and total serum antioxidant capacity with elevation of erythrocyte superoxide dismutase (SOD) and glutathione peroxidase (GPX) levels (Ejtahed et al., 2011).
  • ➢ Bifidobacterium adolescentis improves insulin sensitivity by increased production of glucagon-like peptide 1 (GLP-1) (Chen et al., 2012).
  • ➢ L. acidophilus La5, B. lactis Bb12 decreased fasting blood glucose levels and HbA1c, increased erythrocyte superoxide dismutase, glutathione peroxidase activities, and total antioxidant status (Ejtahed et al., 2012).
  • ➢ L. acidophilus, L. casei, L. bulgaricus, L. bifidum, B. longam, B. breve and s. thermophilus rise in fasting plasma glucose levels and resulted in a decrease in serum hs-CRP and an increase in plasma total GSH (Asemi et al., 2013).
  • ➢ L. reuteri GMNL-263 have been demonstrated to suppress serum glucose, insulin, leptin, C peptide, glycated hemoglobin, GLP-1 level, inflammatory IL-6 and TNF in adipose tissues and PPAR andGLUT4 gene expression in high fructose-fed rats (Hsieh et al., 2013).
  • ➢ L. sporogens had significant effects on serum insulin, hs-CRP, uric acid and plasma total GSH levels (Asemi et al., 2014).
  • ➢ Lb. kefiranofaciens M and Lb. kefiri K prevented the onset of T1D by stimulating the production of GLP-1, regulating the immune-modulatory reaction, and modifying the microbiota (Wang et al., 2012).

A combination of probiotics with the ability to inhibit α-glucosidase and a prebiotic can be used to mitigate T2D and also protect the probiotics from adverse conditions of the GIT.

It is, therefore, possible that an oral supplementation of probiotics with antihyperglycemic properties might be beneficial to T2D patients (Lawrence et al., 2015). The administration of probiotics may improve the prognosis of diabetes through modulation of gut microbiota. Probiotics increase insulin sensitivity and reduce autoimmune response by reducing the inflammatory response and oxidative stress, as well as increasing the expression of adhesion proteins within the intestinal epithelium, reducing intestinal permeability.

6.2 Obesity

  • ➢ L. sporogenes experienced a mean 32% reduction in total cholesterol and 35% reduction in LDL cholesterol over a 3-month period (Mohan, 1990).
  • ➢ L. gasseri, abdominal, visceral, and subcutaneous fat areas decreased significantly. Body weight also decreased significantly (Lu et al., 2004).
  • ➢ S. thermophilus and E. faecium resulted in an 84% reduction in LDL and an increase in fibrinogen levels and precipitation of cholesterol with free bile acids, formed in the media because the activity of the bacterial enzyme bile salt hydrolase (Parvez et al., 2005).
  • ➢ L. paracasei Angiopoietin-related protein 4 (Angptl4), a lipoprotein lipase inhibitor which inhibits the uptake of fatty acids from circulating triglyceride-rich lipoproteins in white adipose and muscle tissues was found to be increased (Aronsson et al., 2010).
  • ➢ L. acidophilus NCFM and L. gasseri SBT2055 showed a decrease in fat mass (Andreasen et al., 2010).
  • ➢ B. pseudocatenulatum SPM 1204, B. longum SPM 1205, and B. longum SPM 1207 has been shown to decrease body weight gain and fat accumulation, blood serum levels of total cholesterol, HDL-C, LDL-C, triglyceride, glucose, leptin, and liver toxicity biomarkers (AST, ALT) (An et al., 2011).
  • ➢ Pediococcus pentosaceus LP28/Lactobacillus plantarum SN13T has been shown to decrease body weight gain, visceral fat accumulation and liver lipid contents (triglyceride and cholesterol) and hepatic lipid droplet accumulation and adipocyte size (Zhao et al., 2012).
  • ➢ Lactobacillus curvatus HY7601 or Lactobacillus curvatus HY7601 in combination with Lactobacillus plantarumKY1032 effectively suppressed body weight gain and reduced the adipose tissue weight in mice fed a high-fat high-cholesterol diet for 9 weeks (Yoo et al., 2013).
  • ➢ Lactobacillus acidophilus, L. bulgaricus, L. bifidum, and L. casei shown to decrease triglyceride, malondialdehyde (MDA), IL-6 and insulin resistance (Mazloom et al., 2013).
  • ➢ Saccharomyces boulardii Biocodex shown to decrease body weight gain and fat mass, hepatic steatosis and total liver lipids content, decreases hepatic (50% decrease in CD11c macrophages level, F4/80, IL-1β and MCP-1mRNA) and systemic inflammation (↓ plasma cytokine concentrations of IL-6, IL-4, IL-1β and TNF-α) (Everard et al., 2014).
  • ➢ Lactobacillus rhamnosus GG shown ↑ glucose tolerance ↑ insulin-stimulated Akt phosphorylation and GLUT4 translocation in skeletal muscle ↓ endoplasmic reticulum (ER) stress in skeletal muscle ↓ M1-like macrophage activation in white adipose tissues ↑ insulin sensitivity (Park et al., 2015).

6.3 Liver Diseases

  • ➢ Lactobacillus casei Normalized phagocytic capacity, decreased TLR4, sTNFR1, sTNFR2, and IL10 levels (Loguercio et al., 2005).
  • ➢ Lactobacillus acidophilus, Lactobacillus helveticus, and Bifidobacterium in rats with alcohol pancreatitis-related liver damage effectively protected against endotoxin/bacterial translocation, as well as liver damage in the course of acute pancreatitis and concomitant heavy alcohol consumption (Marotta et al., 2005).
  • ➢ Lactobacillus, Bifidobacterium and Bacteroides. These changes reduce the expression of TNF-α, IL-1β, and IL-6, and attenuate necroinflammation of the liver (Nardone et al., 2010).
  • ➢ Lactulose versus l-ornithine l-aspartate versus probiotics 110 billion colony-forming units twice daily for 3 months improved minimal hepatic encephalopathy and improve quality of life (Kuczynski et al., 2011).
  • ➢ Lactobacillus rhamnosus, 12 billion CFU/day for 8 weeks, improved transaminases, reduced lipopolysaccharide levels (Vajro et al., 2011).
  • ➢ Lactobacillus rhamnosus GG shown to reduced plasma ALT, endotoxin level, liver steatosis, and inflammation as increasing HIF-mediated mucosal protecting factors and tight junction proteins, positive modification of gut microflora and reduction of endotoxemia (Wang et al., 2011).
  • ➢ L. plantarum encapsulated alginate beads induce a dose-dependent reduction of endotoxin level in rats exposed to alcohol. Also, a reduction in liver function test was observed, as well as molecular markers of inflammation (e.g., NF-κβ, TNF-α and IL-12/p40 subunit) (Murguia-Peniche et al., 2013).
  • ➢ Lactobacillus GG supplementation reduced hepatic inflammation and markedly reduced TNF-expression (Wang et al., 2013).
  • ➢ L. acidophilus and B. longum shown to improve intestinal permeability, attenuated hepatic fat accumulation and butyrate-producing probiotic, Clostridium butyricum, decreased hepatic inflammatory indexes, insulin resistance, triglycerides content, and endotoxin level (Endo et al., 2013).
  • ➢ Bifidobacterium adolescentis SPM0212 found increased expression of myxovirus (Mx) resistance A, decreased extracellular surface antigen of HBV level and the gene expression was inhibited by 40% in HepG2.2.15 cells (Lee et al., 2013).
  • ➢ Reduced risk of hospitalization for HE (hepatic encephalopathy), improved CTP (Child-Turcotte-Pugh) and MELD (model for end-stage liver disease) scores (Dhiman et al., 2014).
  • ➢ Saccharomyces boulardii is able to reduce hepatic steatosis through lowering the hepatic lipid content and low-grade systemic inflammation (Everard et al., 2014).
  • ➢ Lactobacillus rhamnosus R0011 and Lactobacillus Acidophilus acts as modulation of the gut-liver axis: reduction of ALT, TLR4, TNF-, and IL-1 expression, increasing IL-10 expression (Hong et al., 2015).
  • ➢ Lactobacillus rhamnosus and acidophilus mildly decreased intrahepatic lymphocytes and TNF-α expression, as well as reverse irregular and deteriorated microvilli due to alcohol exposure (Hong et al., 2015).

6.4 Cancer

  • ➢ B. longum also plays a role as an immunomodulator and biological response modifier by providing an additional mechanism of tumor suppression. Probiotics also stimulate apoptosis through end-product formation (Okawa et al., 1993).
  • ➢ Bifidobacterium produces lactic acid and lower the intestinal pH and therefore develops a favorable environment which modulates the bacterial enzymes and has cell wall antitumor activities and induces activation of phagocytes to destroy growing tumor cells (Sekine et al., 1994).
  • ➢ Lactobacillus rhamnosus GG, Bifidobacterium lactis Bb12 and oligofructose-enriched inulin reduced proliferation and DNA damage in colonic mucosa and the capacity of fecal water samples to induce necrosis in colonic cells in polypectomized patients. Increased production of interferon (IFN)-γ by peripheral blood mononuclear cells (PBMC) was observed in the cancer patients (Rafter et al., 2007).
  • ➢ Lactobacillus augments the functions of macrophages, natural killer (NK) cells and T cells and exerts anti-cancer activity (Kato et al., 1988).
  • ➢ Bifidobacterium adolescentis SPM1207 had less fecal water content than did control rats, decreasing colon toxicity, due to reduced exposure to soluble toxic compounds (Lee et al., 2009).
  • ➢ Lactobacillus rhamnosus GG administration reduced the number of coliforms and significantly elevated the count of lactobacilli (Bertkova et al., 2010).
  • ➢ L. gasseri OLL2716: LG21 in colorectal cancer patients was shown to decrease alkalosis in stool and cancer markers (fecal product putrescine synthesis oxidized products from incomplete fermentation) (Apás et al., 2010).
  • ➢ B. longum BB536 and L. johnsonii shown probiotic adherence to the colonic mucosa, decreased pathogens and dendritic phenotypes CD83-123, CD83-HLADR, CD83-11c (Gianotti et al., 2010).
  • ➢ Lactobacillus acidophilus and Lactobacillus casei were able to increase the apoptosis-induction capacity of 5-fluorouracil in colorectal carcinoma cell line LS513, suggesting that these probiotics may be used as adjuvants in anticancer chemotherapy (Baldwin et al., 2010).
  • ➢ L. rhamnosus and P. freudenreichii were able to diminish the amount of cancer proteins as c-myc, bcl-2, cyclin D1 and rasp-21 and enhance the levels of GSH, SOD, CAT reduce inflammation and infection (Kumar et al., 2011).
  • ➢ Bifidobacterium or Lactobacillus consumption may limit the formation of toxic metabolites by decreasing the dehydroxylation of primary bile acids and reducing fecal deoxycholic acid concentrations (De Preter et al., 2011).
  • ➢ Propionibacterium freudenreichii increased apoptosis, chromatin condensation, formation of apoptotic bodies, cytotoxicity of camptothecin, DNA laddering, cell cycle arrest and ROS formation (Kumar et al., 2013).

6.5 Diarrhea

6.6 Allergies

Probiotic preparations were also used to treat antibiotic-associated diarrhea, lactose intolerance, dental problems, Helicobacter pylori infections, irritable bowel syndrome, necrotizing enterocolitis, vitamin production, eczema and bacterial vaginosis.

There are several techniques developed to make a required dosage form of probiotics with high efficacy to increase the shelf life of probiotics in its powder form. Some techniques to prepare power form of probiotics are encapsulation, extrusion and emulsification with spray drying, freeze drying, fluidized bed drying and gel bead technology.

Tablets and capsules can be easily designed to control the release, enhancing the adhesion and colonization of probiotic microorganisms to the intestinal epithelium of human host. Several new techniques like PROBIO-TECH, STAR, LIVEBAC, PROBIOCAP, starch encapsulation of LAB, controlled drug delivery, oil matrix complex, cryotabletting, trisphere, and bioadhesive vaginal tablets were developed to provide resistance to stomach acid, to improve stability, to prevent solubilization of probiotics, to release the drug only in intestinal pH, and to improve the shelf life of probiotics (Yadav and Bhitre, 2013).

7 Available Probiotic Food

7.1 Yakult

Yakult is the live (LcS) (Lactobacillus casei Shirota). Yakult contains skimmed milk powder, sugar, glucose, water, and more than 6.5 billion LcS.

Uses:

  1. 1. Prevent digestive disorders such as diarrhea and constipation.
  2. 2. Help build immunity and reduce risk of infections.

7.2 Kefir

Kefir is a microbial symbiotic mixture of lactic acid bacteria (108 CFU/g), yeast (106–107 CFU/g), and acetic acid bacteria (105 CFU/g) that stick to a polysaccharide matrix. The predominant species for kefir preparation are Lactobacillus paracasei ssp. paracasei, Lactobacillus acidophilus, Lactobacillus delbrueckii ssp. bulgaricus, Lactobacillus plantarum, and L. kefiranofaciens.

Uses:

  1. 1. Antitumoral
  2. 2. Anti-inflammatory
  3. 3. Antimicrobial
  4. 4. Immunoregulatory
  5. 5. Antiallergenic
  6. 6. Wound healing
  7. 7. Antidiabetic
  8. 8. Antimutagenic
  9. 9. Antigenotoxic

7.3 Yogurt

Yogurt is a dairy product obtained through the fermentation of milk, partly skimmed milk or skim milk by the lactic bacteria Lactobacillus bulgaricus and Streptococcus thermophilus with which the lactic bacteria Lactobacillus acidophilus and Lactobacilluscasei or the bacteria Bifidobacterium bifidum, Bifidobacterium longum, Bifidobacterium infantis and Bifidobacterium breve may be combined.

Uses:

  1. 1. Prevent osteoporosis
  2. 2. Reduce the risk of high blood pressure
  3. 3. Antibiotic-associated diarrhea and acute diarrhea in children
  4. 4. Vaginal yeast and bacterial infections
  5. 5. Urinary tract infections
  6. 6. Lactose intolerance
  7. 7. Helicobacter pylori infections that cause stomach ulcers
  8. 8. Preventing colorectal cancer

7.4 Kombucha

Is a mixture of black tea and sugar fermentation. Technically, the fermentation becomes a mixture of yeast and bacteria (i.e., Bacterium xylinum, Bacterium gluconicum, Acetobacter ketogenum, and Pichia fermentans).

Uses:

  1. 1. Antidiabetic activity
  2. 2. Reduce atherosclerosis
  3. 3. Antihypertensive effect
  4. 4. Anti-inflammatory
  5. 5. Alleviate arthritis, rheumatism, and gout
  6. 6. Hepatoprotective activity
  7. 7. Cure hemorrhoids
  8. 8. Increase body resistance
  9. 9. Enhance the immune system
  10. 10. Relieve bronchitis and asthma
  11. 11. Reduce menstrual disorders and menopausal hot flashes
  12. 12. Improve hair, skin, and nail health
  13. 13. Reduce stress and nervous disturbances, and insomnia
  14. 14. Improve eyesight

7.5 Sauerkraut

The cabbage is finely shredded, layered with salt, and left to ferment. Fully cured sauerkraut keeps for several months in an airtight container stored at 15°C (60°F) or below by a process of pickling called lactic acid fermentation with Lactobacillus species, including L. brevis and L. plantarum.

Uses:

  1. 1. Increases the bioavailability of nutrients
  2. 2. Promote gut health
  3. 3. Boost circulation
  4. 4. Cardioprotective action
  5. 5. Provide quick energy
  6. 6. Stimulate the immune system
  7. 7. Strengthen bones
  8. 8. Antihyperlipidemic activity
  9. 9. Anti-inflammatory activity
  10. 10. Anticancer property
  11. 11. Improve vision and skin health