Czech Vondrousova, who became the first unseeded woman to lift the title last year, earned a far less welcome accolade after a 6-4, 6-2 defeat by Spain's Jessica Bouzas Maneiro.
"Yeah, it just didn't go as planned today," a downcast Vondrousova said, adding that despite her best efforts the pressure of defending the title had got to her.
"I feel like even if you don't want to think about it, you just think about it... all the time here, I see posters here and everything, my name everywhere.
"Today I was really nervous since the start. I couldn't shake it off. Also, she was playing good. I didn't have many chances to come back into the match or she didn't give me many free points. Yeah, credit to her. I'm going to come back stronger, I hope.
"It was an amazing feeling to go back on Centre Court. Now it's like mixed feelings. I love to be here. I would love to stay longer also. Yeah, today was really tough."
Ranked world number 83, Bouzas Maneiro had won only one tour-level match all year heading into Wimbledon but secured her first win across all Grand Slams when she slapped a backhand down the line.
"It is one of the most important moments of my life," the Spaniard smiled as she soaked up the cheers. "I'm just... I'm surprised with myself to be honest.
"I was just thinking about enjoying the moment and try to play Marketa. She’s one of the best players in the world. She won last year here. I had no pressure, enjoy the moment, enjoy the tournament and just trying to be free-playing and I did it.
"The atmosphere was so nice, so... elegant... it feels like I was at home, I don't know why," she added, explaining her lack of nerves on one of the sport's greatest stages.
The 21-year-old from Galicia hit the ball cleanly throughout an unremarkable Centre Court clash, but only really needed to keep it in play as errors flew from Vondrousova's racket with alarming regularity.
Struggling with a hip injury which derailed her preparation, sixth-seeded Vondrousova now heads for the exit while Bouzas Maneiro will next face compatriot Cristina Bucsa or Romanian Ana Bogdan in the second round.
Disappointment for injured Murray
There was also crushing disappointment for two-time champion Andy Murray.
The British favourite was denied a last hurrah on his beloved Centre Court as his injury curse struck again, the 37-year-old withdrawing from the singles just as the gates opened for day two of the Championships
Murray lost his race against time after surgery to remove a spinal cyst - pulling out of his match against Czech Tomas Machac, although he may still get to write the final chapter of his fabled Wimbledon chronicle with his brother Jamie in the doubles later this week.
While Murray will likely call time on his illustrious career after the Olympics, his old adversary Novak Djokovic showed no sign of his knee problems as he began his quest for a record 25th Grand Slam title impressively.
Serbia's Djokovic had surgery after pulling out of the French Open and was a doubt for Wimbledon but the seven-time champion looked razor sharp as he crushed Czech qualifier Vit Kopriva 6-1 6-2 6-2.
"Practice week went great but an official match on Centre Court is completely different," Djokovic, who risked breaking Wimbledon's all-white clothing policy with a grey knee support, told reporters.
"I didn't know how I would feel or how the knee would hold up, but for an opening match I couldn't ask for a better start."
Light rain disrupted play on a couple of occasions but it did not interfere with women's top seed Iga Swiatek who came through a dangerous first-round against American Sofia Kenin under the Court One roof.
Five-time Grand Slam champion Swiatek was pushed hard by the former Australian Open winner but prevailed 6-3 6-4.
Russian Andrey Rublev became the highest men's seed to depart as the world number six was stunned by Francisco Comesana of Argentina on his Grand Slam debut.
Rublev thumped his racket into his own leg in frustration as he went down 6-4 5-7 6-2 7-6(5).
French Open runner-up Alexander Zverev, the fourth seed, made light work of Spain's Roberto Carballes Baena, while Polish seventh seed Hubert Hurkacz eased past qualifier Radu Albot.
Elena Rybakina, women's champion in 2022, made a bright start as she thrashed qualifier Elena-Gabriela Ruse while there were also wins for American fifth seed Jessica Pegula who dropped two games in dispatching Ashlyn Krueger.