The test sample cholecalciferol (> 98%) was procured from Sigma-Aldrich, India. Similarly, the other chemicals used in the experiments were purchased in India.

The test sample cholecalciferol powder was divided into two parts, i.e., control and treated parts. The control cholecalciferol powder sample did not receive the Trivedi Effect^®-Consciousness Energy Healing Treatment. But, the control cholecalciferol was treated with a “sham” healer who did not have any understanding of the Consciousness Energy Healing Treatment. However, the treated part of cholecalciferol was received the Trivedi Effect^®-Consciousness Energy Healing Treatment remotely under standard laboratory conditions for 3 minutes by the renowned Biofield Energy Healer, Dahryn Trivedi, USA. Finally, both the samples were kept in sealed conditions and characterized using modern analytical techniques.

GC-MS of both the samples of cholecalciferol was analyzed with the help of Perkin Elmer Gas chromatograph equipped with a PE-5MS (30M x 250 micros x 0.250 microns) capillary column and coupled to a single quadrupole mass detector was operated with electron impact (EI) ionization in positive mode. The oven temperature was maintained from 150°C (5 min hold) to 280°C (17 min hold) @ 10°C /min with a total run time of 35 minutes. The sample was prepared taking 50 mg of the cholecalciferol in 2.5 ml methanol as a diluent. The GC-MS based isotopic abundance ratios (P_M+1/P_M and P_M+2/P_M) for the control and Biofield Energy Treated cholecalciferol was calculated using equation (1).

The chromatograms of both the samples are shown in Figure 1. A single major chromatographic peak of the control and treated cholecalciferol was observed at retention time (R_t) of 21.5 and 21.8 minutes, respectively (Figure 1). This indicated that the polarity of both the samples was very close to each other.

The mass spectra of both the samples are shown in Figure 2. The mass spectra of both the samples corresponding to the R_t ~22 minutes exhibited the presence of the molecular ion of cholecalciferol (C₂₇H₄₅O⁺) adduct with hydrogen ion at m/z 385.25 (calcd for C₂₇H₄₅O⁺, 385.35) along with the lower mass peak ^M+at m/z 367.33 (calcd for C₂₇H₄₃⁺, 367.3) (Figure 2 and Figure 3). The experimental data were well matched with the literature data 39.

The cholecalciferol samples showed the mass of a molecular ion at m/z 385.25 (calcd for C₂₇H₄₅O⁺, 385.35)with 100% relative abundance in the spectra. The theoretical calculation of isotopic peak P_M+1 for the protonated cholecalciferol presented as below:

P(13C) = ((27 x 1.1%) x 100% (the actual size of the M⁺ peak)) / 100% = 29.7%

P(2H) = ((45 x 0.015%) x 100%) / 100%= 0.675%

P(17O) = ((1 x 0.04%) x 100%) / 100% = 0.04%

P_M+1i. e.¹³C, ²H, and¹⁷O contributions from C₂₇H₄₅O⁺ to m/z 386.25 = 30.42%

The calculated isotopic abundance of P_M+1 value 30.42% was higher to the experimental value (27.21%) (Table 1). From the above calculation, it has been found that 13C has the major contribution to m/z 386.25.

The LC-MS based isotopic abundance ratio analysis P_M and P_M+1 for cholecalciferol near m/z 385.25 and 386.25, respectively, which were obtained from the observed relative peak intensities of [M⁺] and [(M+1)⁺] peaks, respectively in the ESI-MS spectra (Table 1). The isotopic abundance ratio of P_M+1/P_M (²H/¹H or ¹³C/¹²C or ¹⁷O/¹⁶O) in treated cholecalciferol was increased by 0.74% compared to the control sample (Table 1). Thus, the ¹³C, ²H, and¹⁷O contributions from C₂₇H₄₅O⁺ to m/z 386.25 in the treated cholecalciferol was increased compared to the control sample.

The cholecalciferol samples showed two major independent peaks in the GC-MS chromatograms Figure 3. The Rt of the control cholecalciferol was at 22.06 and 22.74 minutes, whereas 21.95 and 22.62 minutes is for the treated cholecalciferol, which indicated that the polarity of both the sample was very close. The two peaks in the chromatograms of both the cholecalciferol samples might be due to the cis and trans isomers of cholecalciferol 4041.

The GC-MS spectra of the cholecalciferol samples at R_t of 22 minutes exhibited the presence of the molecular ion peak of cholecalciferol (C₂₇H₄₄O⁺) (Figure 4) at m/z 385 (calcd for C₂₇H₄₄O⁺, 384.34). The low molecular mass fragmentation peak at m/z 367, and 352 for C₂₇H₄₃⁺ and C₂₆H₄₀⁺, respectively were also observed in both the spectra (Figure 4). The mass peak intensities of the Biofield Energy Treated cholecalciferol were altered compared to the control sample.

The GC-MS spectra of both the cholecalciferol showed the mass of the molecular ion peak ^M+ at m/z 385 (calcd for C₂₇H₄₄O⁺, 384.34). The theoretical calculation of P_M+1 and P_M+2 for cholecalciferol was presented as below:

P(¹³C) = ((27 x 1.1%) x 10% (the actual size of the M⁺ peak)) / 100% = 2.97%

P(²H) = ((44 x 0.015%) x 10%) / 100%= 0.07%

P(¹⁷O) = ((1 x 0.04%) x 10%) / 100% = 0.004%

P_M+1i. e.¹³C, ²H, and¹⁷O contributions from C₂₇H₄₅O⁺ to m/z 386 = 3.04%

Similarly, the theoretical calculation of isotopic peak P_M+2 for the protonated cholecalciferol was presented below:

P(¹⁸O) = ((1 x 0.20%) x 10%) / 100% = 0.02%

P_M+2 of ¹⁸O contribution from C₂₇H₄₅O⁺ to m/z 387 = 0.02%

The calculated isotopic abundance of P_M+1 and P_M+2 values were close to the calculated value (Table 2). From the above calculation, it has been found that ¹³C and ¹⁸O have major contribution to m/z 386 and 387 of cholecalciferol.

The GC-MS based isotopic abundance ratio analysis of the treated sample was calculated compared to the control sample. P_M, P_M+1, and P_M+2 for cholecalciferol near m/z 385, 386, and 387, respectively of the control and treated samples, which were obtained from the observed relative peak intensities of [M⁺], [(M+1)⁺], and [(M+2)⁺] peaks, respectively in the mass spectra. The isotopic abundance ratio of P_M+1/P_M and P_M+2/P_M in the treated cholecalciferol was significantly increased by 66.39% and 62.69%, respectively compared to the control sample (Table 2). Therefore, the¹³C, ²H, ¹⁷O and 18O contributions from C₂₇H₄₄O⁺ to m/z 386 and 387 in the treated cholecalciferol were significantly increased compared with the control sample. Figure 5.

LC-MS and GC-MS study confirmed the structure of cholecalciferol. The isotopic abundance ratios of P_M+1/P_M (²H/¹H or ¹³C/¹²C or ¹⁷O/¹⁶O) and P_M+2/P_M (¹⁸O/¹⁶O) in the treated cholecalciferol were significantly increased compared to the control sample. The altered isotopic composition of the Trivedi Effect^®-Consciousness Energy Healing Treated cholecalciferol might have altered the neutron to proton ratio in the nucleus via the possible mediation of neutrino 15. Neutrino is a subatomic particle but has no electrical charge and a very small mass. These are one of the most abundant particles in the universe. The neutrinos have the ability to interact with protons and neutrons in the nucleus, which might have a close relation between neutrino and the isotope formation 153435. The isotopic abundance ratios ²H/¹H or ¹³C/¹²C or ¹⁷O/¹⁶O or ¹⁸O/¹⁶O would influence the atomic bond vibration of treated cholecalciferol 42. The increased isotopic abundance ratio of the treated cholecalciferol may increase the intra-atomic bond strength, increase its stability. The Biofield EnergyTreated cholecalciferol would be more stable and suitable for the prevention and treatment of various diseases such as vitamin D deficiency, rickets, osteoporosis, arthritis, multiple sclerosis, cancer, diabetes mellitus, mental disorders, cardiovascular diseases, hypertension, infections, influenza, cognitive impairment in older adults, Parkinson’s and Alzheimer’s diseases, autoimmune disease, dementia, glucose intolerance, multiple sclerosis, etc.