Assessment of Human Carotenoid Status Using Raman Spectroscopy Presented by



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Assessment of Human Carotenoid Status Using Raman Spectroscopy

  • Presented by:

  • Angela Mastaloudis, Ph.D.

  • Pharmanex Research Institute

  • Provo, UT, USA


Carotenoids

  • An important group of phytonutrients

  • Abundant in fruits and vegetables

  • Epidemiological and clinical studies indicate protection against the development of a variety of chronic diseases

    • Lutein and zeaxanthin
      • eye health
    • -carotene and lycopene
      • protection from sun damage
    • Lutein and lycopene
      • cardiovascular health
    • Lycopene
      • prostate cancer


Carotenoids



Relevance of Accurate Assessment of Human Carotenoid Status

  • Marker of oxidative stress

    • smoking, sunlight exposure, pollution all influence skin carotenoids independent of dietary intake
  • Survey fruit and vegetable consumption

  • Monitor compliance to diets rich in fruits and vegetables

  • Monitor intake of carotenoid containing dietary supplements



Resonant Raman Scattering of Carotenoids



Resonant Raman Scattering of Carotenoids



Early Studies: The Eye & AMD



BioPhotonic Scanner

  • BioPhotonic Scanner

  • 473 nm excitation, 511 nm detection (C=C)



Skin Carotenoids

  • Carotenoids are measured in the stratum corneum layer of the skin

  • Carotenoids protect epidermis from UV Light

  • Site of action advantage



Skin Carotenoids

  • An Important Indicator of the Body’s Antioxidant Defense System

  • Most convenient measure of antioxidant status

  • Carotenoids measured:

    • -Carotene, -Carotene, Lycopene, Lutein, Zeaxanthin, -Cryptoxanthin
    • First line of defense & part of antioxidant network
  • Skin carotenoids are influenced by oxidative stress (smoking, sunlight exposure, pollution) independent of dietary intake



Advantages

  • Non-invasive

  • Safe

  • Instant results

  • Inexpensive Highly specific

  • Reflects long-term carotenoid status



Clinical validation of a non-invasive, Raman spectroscopic method to assess carotenoid nutritional status in humans

  • Jeffrey A. Zidichouski1,3, Angela Mastaloudis1, Stephen J. Poole1, James C. Reading2 and Carsten R. Smidt1

  • 1Pharmanex Research Institute, Provo, Utah, USA; 2U. of Utah School of Medicine, SLC, Utah, USA; 3Institute for Nutrisciences and Health, NRC, Canada



Background

  • Serum/plasma carotenoids (HPLC)

    • currently accepted “gold standard”
    • used to validate subjective fruit & vegetable intake data (epidemiological research)
    • used to assess human carotenoid and antioxidant status
    • invasive & labor-intensive
    • not suitable for large populations


Objectives

  • To validate the Raman spectroscopy (RS) methodology (BioPhotonic Scanner) by comparing it to HPLC, the currently accepted gold standard for assessing human carotenoid status using Criterion Validity

  • To assess and compare reliability of the two measures



Methodology

  • 372 adult non-smokers

  • 3 measurements within 8 days, ≥ 48 h apart, after overnight (12 h) fast:

    • Serum carotenoids by HPLC
    • Skin carotenoids, Raman spectroscopy
    • Demographic data and food frequency questionnaires


Results

  • Carotenoids (means ± SD)



Skin-Serum Correlation

  • R = 0.81; p < 0.001



Conclusions

  • Raman spectroscopy (BioPhotonic Scanner) is a valid measure to assess skin carotenoid status in situ in humans

  • RS accurately predicts blood serum total carotenoids, independent of demographic and lifestyle factors

  • Skin carotenoids are more stable over time than serum carotenoids, likely because they are not confounded by recent dietary intake (Dueker SR et al. J Lipid Res. 2000;41:1790)

  • RS appears to be a better indicator of carotenoid status than blood carotenoids



Epidemiological Studies





Fruit & Vegetable Intake



Supplementation: LP* *Multivitamin/mineral Supplement with Antioxidant Nutrients



Body Mass Index



Smoking (Oxidative Stress)





Epidemiological Studies Summary

  • Skin carotenoids:

    • Are not influenced by age, gender or race
    • Increase linearly in response to both dietary intake and intake from supplements
    • Are inversely correlated with oxidative stress markers
      • (smoking, UV light exposure, urinary F2-Isoprostanes)
    • Are a valid indicator of overall antioxidant status


Randomized, Placebo Controlled Antioxidant Supplementation Study



Study Design

  • N = 52

  • RCT: Twice a day, with breakfast and with dinner, subjects took LP nano or Placebo

  • Study duration: 18 weeks

  • BioPhotonic Scanner

    • Subjects were scanned a total of 15 times
  • Lifestyle





Summary

  • Skin carotenoids increased significantly with supplementation ( two weeks)

  • Skin carotenoids increased in all 20 subjects in twenty weeks with supplementation

  • Even after 18 weeks, skin carotenoids were still increasing with supplementation



Double-Blind, Placebo-Controlled Antioxidant Supplementation Study

  • RLI, Florida Hospital Celebration Health

  • 53 subjects who had not taken antioxidant supplements within the last 3 months

  • 6 weeks on supplements (LP or Placebo)

  • Measured skin RS response and serum antioxidants at days 0, 21 and 42



Double-Blind, Placebo-Controlled Antioxidant Supplementation Study



Results: Serum Carotenoids



Results: Skin Carotenoids



Results: Serum Vitamins C and E



Can Skin Carotenoids Serve as a Marker of Antioxidant Status or Oxidative Stress?



The Antioxidant Network



Associations of Antioxidant Status and Oxidative Stress with Skin Carotenoids Assessed by Raman Spectroscopy

  • Joseph Carlson1,3, Shayn Stavens1,

  • Richard Holubkav1, Jeffrey Zidichouski2,

  • Angela Mastaloudis2, Carsten Smidt2, Eldon Askew1

  • 1U. of Utah Salt Lake City, Utah; 2Pharmanex Research Institute Provo, UT; 3Michigan State U. East Lansing, MI



Objectives

  • To evaluate the relationship between skin carotenoids and serum antioxidants (vitamins E and C)

  • To evaluate the relationship between carotenoid antioxidant status and markers of oxidative stress



Study Design

  • N = 307 adult non-smokers

  • Serum carotenoids, vitamin E and vitamin C by HPLC

  • Skin carotenoids using Raman spectroscopy

  • Questionnaires: demographic, lifestyle and dietary data

  • Urinary F2-Isoprostanes



F2-Isoprostanes

  • Gold standard for measurement of in vivo lipid peroxidation

  • Reliable

  • Chemically stable

  • Specific end-product of the free radical mediated oxidation of arachidonic acid (20:4)

  • Possess biological pro-atherogenic activity

  • Plasma concentrations are elevated in subjects with known oxidative stress (smokers, diabetics, obesity, cardiovascular disease, Alzheimer’s Disease etc)



Skin and Serum Carotenoids are Strongly Correlated



Skin Carotenoids are Positively Correlated with Plasma Antioxidants



Skin Carotenoids are Inversely Correlated with F2-Isoprostanes



Summary

  • Skin carotenoids assessed by the Biophotonic Scanner were directly correlated with:

    • Serum Carotenoids
    • Plasma antioxidants (Vitamins E and C)
    • Fruit and vegetable intake
  • Skin carotenoids were inversely related to F2-Isoprosanes, a marker of oxidative stress



Taiwan Data (n = 38,843; April 2006)



Taiwan Data (n = 38,942; April 2006)



Taiwan Data (n = 38,907; April 2006)



Conclusions

  • Measurement of skin carotenoids using Raman spectroscopy (BioPhotonic Scanner) is:

    • highly correlated with serum carotenoids
    • less variable than serum carotenoids
    • preferable to serum carotenoids as a marker of fruit and vegetable intake
    • Reflective of long-term carotenoid status
    • indicative of other serum antioxidants (vitamins C and E)
    • indicative of oxidative stress (MDA,F2-Isoprostanes)


Summary

  • Resonance Raman scattering is a viable optical technique to measure skin carotenoids in vivo. It is highly specific, non-invasive and suitable for clinical field measurements of large populations.



Acknowledgments

  • Stephen Poole, Carsten Smidt – Pharmanex Research Institute

  • Jason Morrow – Vanderbilt University

  • Neal Craft – Craft Technologies, NC

  • Lester Packer – University of Southern California

  • Kyung-Jin Yeum, Jeff Blumberg – Tufts University

  • James Rippe – Rippe Lifestyle Institute

  • Wayne Askew, Joe Carlson, Shayn Stavens

    • University of Utah, Division of Foods and Nutrition, SLC
  • Jeff Zidichouski – Canadian National Research Council (P.E.I.)

  • Werner Gellermann – University of Utah, Physics Dept., SLC



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